CN117599181A - Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms - Google Patents
Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms Download PDFInfo
- Publication number
- CN117599181A CN117599181A CN202311588201.3A CN202311588201A CN117599181A CN 117599181 A CN117599181 A CN 117599181A CN 202311588201 A CN202311588201 A CN 202311588201A CN 117599181 A CN117599181 A CN 117599181A
- Authority
- CN
- China
- Prior art keywords
- pain
- chronic pain
- vta
- mice
- injection
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
- 208000002193 Pain Diseases 0.000 title claims abstract description 153
- 208000000094 Chronic Pain Diseases 0.000 title claims abstract description 107
- 208000024891 symptom Diseases 0.000 title claims abstract description 97
- 210000002569 neuron Anatomy 0.000 title claims abstract description 34
- 238000002360 preparation method Methods 0.000 title claims abstract description 14
- 239000003814 drug Substances 0.000 claims abstract description 55
- 210000001362 glutamatergic neuron Anatomy 0.000 claims abstract description 44
- BTCSSZJGUNDROE-UHFFFAOYSA-N gamma-aminobutyric acid Chemical compound NCCCC(O)=O BTCSSZJGUNDROE-UHFFFAOYSA-N 0.000 claims abstract description 26
- 206010063659 Aversion Diseases 0.000 claims abstract description 25
- 208000019901 Anxiety disease Diseases 0.000 claims abstract description 24
- 238000011282 treatment Methods 0.000 claims abstract description 24
- 230000036506 anxiety Effects 0.000 claims abstract description 17
- OGNSCSPNOLGXSM-UHFFFAOYSA-N (+/-)-DABA Natural products NCCC(N)C(O)=O OGNSCSPNOLGXSM-UHFFFAOYSA-N 0.000 claims abstract description 13
- 229960003692 gamma aminobutyric acid Drugs 0.000 claims abstract description 13
- 208000010877 cognitive disease Diseases 0.000 claims abstract description 10
- 208000028698 Cognitive impairment Diseases 0.000 claims abstract description 9
- 210000004326 gyrus cinguli Anatomy 0.000 claims abstract description 6
- 102000004980 Dopamine D2 Receptors Human genes 0.000 claims abstract description 5
- 108090001111 Dopamine D2 Receptors Proteins 0.000 claims abstract description 5
- 210000004515 ventral tegmental area Anatomy 0.000 claims description 118
- 239000007924 injection Substances 0.000 claims description 79
- 238000002347 injection Methods 0.000 claims description 79
- 230000000694 effects Effects 0.000 claims description 74
- 210000005064 dopaminergic neuron Anatomy 0.000 claims description 67
- LNEPOXFFQSENCJ-UHFFFAOYSA-N haloperidol Chemical group C1CC(O)(C=2C=CC(Cl)=CC=2)CCN1CCCC(=O)C1=CC=C(F)C=C1 LNEPOXFFQSENCJ-UHFFFAOYSA-N 0.000 claims description 58
- 101150049660 DRD2 gene Proteins 0.000 claims description 54
- 230000036407 pain Effects 0.000 claims description 45
- 229960003878 haloperidol Drugs 0.000 claims description 29
- 239000000843 powder Substances 0.000 claims description 23
- 230000001537 neural effect Effects 0.000 claims description 20
- 229940044601 receptor agonist Drugs 0.000 claims description 20
- 239000000018 receptor agonist Substances 0.000 claims description 20
- 230000000848 glutamatergic effect Effects 0.000 claims description 18
- FTSUPYGMFAPCFZ-ZWNOBZJWSA-N quinpirole Chemical group C([C@H]1CCCN([C@@H]1C1)CCC)C2=C1C=NN2 FTSUPYGMFAPCFZ-ZWNOBZJWSA-N 0.000 claims description 18
- 229950001037 quinpirole Drugs 0.000 claims description 18
- 229940044551 receptor antagonist Drugs 0.000 claims description 18
- 239000002464 receptor antagonist Substances 0.000 claims description 18
- 229940079593 drug Drugs 0.000 claims description 16
- 230000002401 inhibitory effect Effects 0.000 claims description 16
- 210000001222 gaba-ergic neuron Anatomy 0.000 claims description 15
- -1 elixir Substances 0.000 claims description 9
- 239000003112 inhibitor Substances 0.000 claims description 9
- 208000019695 Migraine disease Diseases 0.000 claims description 8
- 208000008765 Sciatica Diseases 0.000 claims description 8
- 150000001875 compounds Chemical class 0.000 claims description 8
- 229910052736 halogen Inorganic materials 0.000 claims description 8
- 150000002367 halogens Chemical group 0.000 claims description 8
- 238000004519 manufacturing process Methods 0.000 claims description 8
- 206010027599 migraine Diseases 0.000 claims description 8
- 208000004296 neuralgia Diseases 0.000 claims description 8
- 208000021722 neuropathic pain Diseases 0.000 claims description 8
- 150000003839 salts Chemical class 0.000 claims description 8
- 206010044652 trigeminal neuralgia Diseases 0.000 claims description 8
- 239000008194 pharmaceutical composition Substances 0.000 claims description 7
- 239000000243 solution Substances 0.000 claims description 7
- 239000002775 capsule Substances 0.000 claims description 6
- 239000006071 cream Substances 0.000 claims description 6
- 239000003623 enhancer Substances 0.000 claims description 6
- 238000009472 formulation Methods 0.000 claims description 6
- 239000008187 granular material Substances 0.000 claims description 6
- 239000012669 liquid formulation Substances 0.000 claims description 6
- 239000000203 mixture Substances 0.000 claims description 6
- 239000002674 ointment Substances 0.000 claims description 6
- 239000007787 solid Substances 0.000 claims description 6
- 239000000725 suspension Substances 0.000 claims description 6
- 230000003920 cognitive function Effects 0.000 claims description 5
- 230000006735 deficit Effects 0.000 claims description 5
- 125000006273 (C1-C3) alkyl group Chemical group 0.000 claims description 4
- 125000004169 (C1-C6) alkyl group Chemical group 0.000 claims description 4
- 230000002708 enhancing effect Effects 0.000 claims description 4
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 claims description 4
- KORNTPPJEAJQIU-KJXAQDMKSA-N Cabaser Chemical compound C1=CC([C@H]2C[C@H](CN(CC=C)[C@@H]2C2)C(=O)N(CCCN(C)C)C(=O)NCC)=C3C2=CNC3=C1 KORNTPPJEAJQIU-KJXAQDMKSA-N 0.000 claims description 3
- 229960002802 bromocriptine Drugs 0.000 claims description 3
- OZVBMTJYIDMWIL-AYFBDAFISA-N bromocriptine Chemical compound C1=CC(C=2[C@H](N(C)C[C@@H](C=2)C(=O)N[C@]2(C(=O)N3[C@H](C(N4CCC[C@H]4[C@]3(O)O2)=O)CC(C)C)C(C)C)C2)=C3C2=C(Br)NC3=C1 OZVBMTJYIDMWIL-AYFBDAFISA-N 0.000 claims description 3
- 229960004596 cabergoline Drugs 0.000 claims description 3
- 229960004851 pergolide Drugs 0.000 claims description 3
- YEHCICAEULNIGD-MZMPZRCHSA-N pergolide Chemical compound C1=CC([C@H]2C[C@@H](CSC)CN([C@@H]2C2)CCC)=C3C2=CNC3=C1 YEHCICAEULNIGD-MZMPZRCHSA-N 0.000 claims description 3
- 229960003089 pramipexole Drugs 0.000 claims description 3
- FASDKYOPVNHBLU-ZETCQYMHSA-N pramipexole Chemical compound C1[C@@H](NCCC)CCC2=C1SC(N)=N2 FASDKYOPVNHBLU-ZETCQYMHSA-N 0.000 claims description 3
- BGRJTUBHPOOWDU-NSHDSACASA-N (S)-(-)-sulpiride Chemical compound CCN1CCC[C@H]1CNC(=O)C1=CC(S(N)(=O)=O)=CC=C1OC BGRJTUBHPOOWDU-NSHDSACASA-N 0.000 claims description 2
- QCQCHGYLTSGIGX-GHXANHINSA-N 4-[[(3ar,5ar,5br,7ar,9s,11ar,11br,13as)-5a,5b,8,8,11a-pentamethyl-3a-[(5-methylpyridine-3-carbonyl)amino]-2-oxo-1-propan-2-yl-4,5,6,7,7a,9,10,11,11b,12,13,13a-dodecahydro-3h-cyclopenta[a]chrysen-9-yl]oxy]-2,2-dimethyl-4-oxobutanoic acid Chemical compound N([C@@]12CC[C@@]3(C)[C@]4(C)CC[C@H]5C(C)(C)[C@@H](OC(=O)CC(C)(C)C(O)=O)CC[C@]5(C)[C@H]4CC[C@@H]3C1=C(C(C2)=O)C(C)C)C(=O)C1=CN=CC(C)=C1 QCQCHGYLTSGIGX-GHXANHINSA-N 0.000 claims description 2
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical compound [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 claims description 2
- PXGOKWXKJXAPGV-UHFFFAOYSA-N Fluorine Chemical compound FF PXGOKWXKJXAPGV-UHFFFAOYSA-N 0.000 claims description 2
- 229910052801 chlorine Inorganic materials 0.000 claims description 2
- 239000000460 chlorine Substances 0.000 claims description 2
- 229960001253 domperidone Drugs 0.000 claims description 2
- FGXWKSZFVQUSTL-UHFFFAOYSA-N domperidone Chemical compound C12=CC=CC=C2NC(=O)N1CCCN(CC1)CCC1N1C2=CC=C(Cl)C=C2NC1=O FGXWKSZFVQUSTL-UHFFFAOYSA-N 0.000 claims description 2
- 229910052731 fluorine Inorganic materials 0.000 claims description 2
- 239000011737 fluorine Substances 0.000 claims description 2
- 229960004503 metoclopramide Drugs 0.000 claims description 2
- TTWJBBZEZQICBI-UHFFFAOYSA-N metoclopramide Chemical compound CCN(CC)CCNC(=O)C1=CC(Cl)=C(N)C=C1OC TTWJBBZEZQICBI-UHFFFAOYSA-N 0.000 claims description 2
- 229960004940 sulpiride Drugs 0.000 claims description 2
- 206010045171 Tumour pain Diseases 0.000 claims 4
- 208000018737 Parkinson disease Diseases 0.000 claims 3
- VYFYYTLLBUKUHU-UHFFFAOYSA-N dopamine Chemical compound NCCC1=CC=C(O)C(O)=C1 VYFYYTLLBUKUHU-UHFFFAOYSA-N 0.000 abstract description 10
- 229960003638 dopamine Drugs 0.000 abstract description 5
- 241000699670 Mus sp. Species 0.000 description 116
- 229940090044 injection Drugs 0.000 description 60
- 238000002474 experimental method Methods 0.000 description 55
- 238000012360 testing method Methods 0.000 description 43
- 238000010172 mouse model Methods 0.000 description 32
- 241000700605 Viruses Species 0.000 description 29
- 210000004556 brain Anatomy 0.000 description 27
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 24
- 230000006399 behavior Effects 0.000 description 22
- 238000011156 evaluation Methods 0.000 description 19
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 18
- 239000002504 physiological saline solution Substances 0.000 description 18
- 230000036982 action potential Effects 0.000 description 16
- 230000003040 nociceptive effect Effects 0.000 description 15
- 230000007246 mechanism Effects 0.000 description 14
- 230000003542 behavioural effect Effects 0.000 description 13
- 239000000523 sample Substances 0.000 description 13
- 239000011550 stock solution Substances 0.000 description 13
- 230000005856 abnormality Effects 0.000 description 12
- 241000699666 Mus <mouse, genus> Species 0.000 description 11
- 210000004027 cell Anatomy 0.000 description 11
- QZUDBNBUXVUHMW-UHFFFAOYSA-N clozapine Chemical compound C1CN(C)CCN1C1=NC2=CC(Cl)=CC=C2NC2=CC=CC=C12 QZUDBNBUXVUHMW-UHFFFAOYSA-N 0.000 description 11
- 230000000638 stimulation Effects 0.000 description 11
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 9
- 230000004913 activation Effects 0.000 description 9
- 230000007529 anxiety like behavior Effects 0.000 description 9
- 230000006397 emotional response Effects 0.000 description 9
- 230000004044 response Effects 0.000 description 9
- 241001465754 Metazoa Species 0.000 description 8
- 238000011161 development Methods 0.000 description 8
- 210000005036 nerve Anatomy 0.000 description 8
- 239000011780 sodium chloride Substances 0.000 description 8
- 210000001175 cerebrospinal fluid Anatomy 0.000 description 7
- 229930006000 Sucrose Natural products 0.000 description 6
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 6
- 239000000556 agonist Substances 0.000 description 6
- 239000005557 antagonist Substances 0.000 description 6
- 229960004170 clozapine Drugs 0.000 description 6
- 230000001149 cognitive effect Effects 0.000 description 6
- 230000007774 longterm Effects 0.000 description 6
- 238000000034 method Methods 0.000 description 6
- 230000001105 regulatory effect Effects 0.000 description 6
- 239000005720 sucrose Substances 0.000 description 6
- 230000007423 decrease Effects 0.000 description 5
- 238000012377 drug delivery Methods 0.000 description 5
- 230000002829 reductive effect Effects 0.000 description 5
- 238000011200 topical administration Methods 0.000 description 5
- 208000000114 Pain Threshold Diseases 0.000 description 4
- 208000027089 Parkinsonian disease Diseases 0.000 description 4
- 206010034010 Parkinsonism Diseases 0.000 description 4
- 230000003213 activating effect Effects 0.000 description 4
- 238000001514 detection method Methods 0.000 description 4
- 230000007831 electrophysiology Effects 0.000 description 4
- 238000002001 electrophysiology Methods 0.000 description 4
- 230000008451 emotion Effects 0.000 description 4
- 230000001965 increasing effect Effects 0.000 description 4
- 230000000771 oncological effect Effects 0.000 description 4
- 230000008533 pain sensitivity Effects 0.000 description 4
- 230000037040 pain threshold Effects 0.000 description 4
- 210000003497 sciatic nerve Anatomy 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 230000002459 sustained effect Effects 0.000 description 4
- 210000001519 tissue Anatomy 0.000 description 4
- 229920000049 Carbon (fiber) Polymers 0.000 description 3
- PIWKPBJCKXDKJR-UHFFFAOYSA-N Isoflurane Chemical compound FC(F)OC(Cl)C(F)(F)F PIWKPBJCKXDKJR-UHFFFAOYSA-N 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 230000033228 biological regulation Effects 0.000 description 3
- 239000004917 carbon fiber Substances 0.000 description 3
- 230000019771 cognition Effects 0.000 description 3
- 230000007267 depressive like behavior Effects 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 230000003291 dopaminomimetic effect Effects 0.000 description 3
- 238000010304 firing Methods 0.000 description 3
- 230000006870 function Effects 0.000 description 3
- 238000001727 in vivo Methods 0.000 description 3
- 229960002725 isoflurane Drugs 0.000 description 3
- 230000013016 learning Effects 0.000 description 3
- 238000012423 maintenance Methods 0.000 description 3
- 230000001404 mediated effect Effects 0.000 description 3
- VNWKTOKETHGBQD-UHFFFAOYSA-N methane Chemical compound C VNWKTOKETHGBQD-UHFFFAOYSA-N 0.000 description 3
- 230000036651 mood Effects 0.000 description 3
- 230000008587 neuronal excitability Effects 0.000 description 3
- 230000003287 optical effect Effects 0.000 description 3
- 239000013307 optical fiber Substances 0.000 description 3
- 230000001225 therapeutic effect Effects 0.000 description 3
- 206010013954 Dysphoria Diseases 0.000 description 2
- WHUUTDBJXJRKMK-VKHMYHEASA-N L-glutamic acid Chemical compound OC(=O)[C@@H](N)CCC(O)=O WHUUTDBJXJRKMK-VKHMYHEASA-N 0.000 description 2
- 229920005372 Plexiglas® Polymers 0.000 description 2
- 208000027418 Wounds and injury Diseases 0.000 description 2
- 230000004075 alteration Effects 0.000 description 2
- 230000003930 cognitive ability Effects 0.000 description 2
- 230000006378 damage Effects 0.000 description 2
- 230000000994 depressogenic effect Effects 0.000 description 2
- 208000035475 disorder Diseases 0.000 description 2
- 238000004043 dyeing Methods 0.000 description 2
- 230000002996 emotional effect Effects 0.000 description 2
- 239000000835 fiber Substances 0.000 description 2
- 229930195712 glutamate Natural products 0.000 description 2
- 229940047551 haloperidol injection Drugs 0.000 description 2
- 230000005764 inhibitory process Effects 0.000 description 2
- 208000014674 injury Diseases 0.000 description 2
- 238000007917 intracranial administration Methods 0.000 description 2
- 239000007928 intraperitoneal injection Substances 0.000 description 2
- 210000003141 lower extremity Anatomy 0.000 description 2
- 230000008534 mechanical pain sensitivity Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 230000008058 pain sensation Effects 0.000 description 2
- 238000012402 patch clamp technique Methods 0.000 description 2
- 230000002085 persistent effect Effects 0.000 description 2
- 239000004033 plastic Substances 0.000 description 2
- 229920003023 plastic Polymers 0.000 description 2
- 239000004926 polymethyl methacrylate Substances 0.000 description 2
- 230000001242 postsynaptic effect Effects 0.000 description 2
- 230000000750 progressive effect Effects 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 230000001953 sensory effect Effects 0.000 description 2
- 230000020341 sensory perception of pain Effects 0.000 description 2
- 230000004936 stimulating effect Effects 0.000 description 2
- 210000001590 sural nerve Anatomy 0.000 description 2
- 230000005062 synaptic transmission Effects 0.000 description 2
- 210000002972 tibial nerve Anatomy 0.000 description 2
- 230000003612 virological effect Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 206010001497 Agitation Diseases 0.000 description 1
- 206010002091 Anaesthesia Diseases 0.000 description 1
- 244000025254 Cannabis sativa Species 0.000 description 1
- 229940096895 Dopamine D2 receptor agonist Drugs 0.000 description 1
- 229940123603 Dopamine D2 receptor antagonist Drugs 0.000 description 1
- 208000012661 Dyskinesia Diseases 0.000 description 1
- 239000008896 Opium Substances 0.000 description 1
- 229930040373 Paraformaldehyde Natural products 0.000 description 1
- 206010033799 Paralysis Diseases 0.000 description 1
- 238000000692 Student's t-test Methods 0.000 description 1
- 230000002159 abnormal effect Effects 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 230000037005 anaesthesia Effects 0.000 description 1
- 229940035676 analgesics Drugs 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 239000000730 antalgic agent Substances 0.000 description 1
- 230000006400 anxiety behaviour Effects 0.000 description 1
- 210000003050 axon Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 238000004061 bleaching Methods 0.000 description 1
- 238000009933 burial Methods 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 208000022371 chronic pain syndrome Diseases 0.000 description 1
- 229940121657 clinical drug Drugs 0.000 description 1
- 238000007596 consolidation process Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 239000000442 dopamine 2 receptor blocking agent Substances 0.000 description 1
- 239000002288 dopamine 2 receptor stimulating agent Substances 0.000 description 1
- 239000003596 drug target Substances 0.000 description 1
- 230000008918 emotional behaviour Effects 0.000 description 1
- 230000000763 evoking effect Effects 0.000 description 1
- 210000003414 extremity Anatomy 0.000 description 1
- 238000000556 factor analysis Methods 0.000 description 1
- 210000002683 foot Anatomy 0.000 description 1
- 230000035876 healing Effects 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 231100000206 health hazard Toxicity 0.000 description 1
- 210000000548 hind-foot Anatomy 0.000 description 1
- 238000003384 imaging method Methods 0.000 description 1
- 230000002055 immunohistochemical effect Effects 0.000 description 1
- 238000003364 immunohistochemistry Methods 0.000 description 1
- 238000012744 immunostaining Methods 0.000 description 1
- 210000005240 left ventricle Anatomy 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 229910052751 metal Inorganic materials 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 238000000520 microinjection Methods 0.000 description 1
- 230000009456 molecular mechanism Effects 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 230000007230 neural mechanism Effects 0.000 description 1
- 230000008906 neuronal response Effects 0.000 description 1
- 229960001027 opium Drugs 0.000 description 1
- 230000037324 pain perception Effects 0.000 description 1
- 238000002559 palpation Methods 0.000 description 1
- 229920002866 paraformaldehyde Polymers 0.000 description 1
- 210000004345 peroneal nerve Anatomy 0.000 description 1
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 230000009024 positive feedback mechanism Effects 0.000 description 1
- 230000010411 postconditioning Effects 0.000 description 1
- 230000002980 postoperative effect Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 230000007425 progressive decline Effects 0.000 description 1
- 108020003175 receptors Proteins 0.000 description 1
- 102000005962 receptors Human genes 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000011514 reflex Effects 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 210000005241 right ventricle Anatomy 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 230000035807 sensation Effects 0.000 description 1
- 231100000933 sensitization response Toxicity 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 208000019116 sleep disease Diseases 0.000 description 1
- 230000002269 spontaneous effect Effects 0.000 description 1
- 229910001220 stainless steel Inorganic materials 0.000 description 1
- 239000010935 stainless steel Substances 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 238000001356 surgical procedure Methods 0.000 description 1
- 230000009182 swimming Effects 0.000 description 1
- 230000000946 synaptic effect Effects 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 230000008536 thermal pain sensitivity Effects 0.000 description 1
- 210000000689 upper leg Anatomy 0.000 description 1
- 230000002618 waking effect Effects 0.000 description 1
- 230000003313 weakening effect Effects 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/16—Amides, e.g. hydroxamic acids
- A61K31/165—Amides, e.g. hydroxamic acids having aromatic rings, e.g. colchicine, atenolol, progabide
- A61K31/166—Amides, e.g. hydroxamic acids having aromatic rings, e.g. colchicine, atenolol, progabide having the carbon of a carboxamide group directly attached to the aromatic ring, e.g. procainamide, procarbazine, metoclopramide, labetalol
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/40—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having five-membered rings with one nitrogen as the only ring hetero atom, e.g. sulpiride, succinimide, tolmetin, buflomedil
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/44—Non condensed pyridines; Hydrogenated derivatives thereof
- A61K31/445—Non condensed piperidines, e.g. piperocaine
- A61K31/4523—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems
- A61K31/454—Non condensed piperidines, e.g. piperocaine containing further heterocyclic ring systems containing a five-membered ring with nitrogen as a ring hetero atom, e.g. pimozide, domperidone
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/47—Quinolines; Isoquinolines
- A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
- A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/33—Heterocyclic compounds
- A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
- A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
- A61K31/47—Quinolines; Isoquinolines
- A61K31/48—Ergoline derivatives, e.g. lysergic acid, ergotamine
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/02—Drugs for disorders of the nervous system for peripheral neuropathies
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/06—Antimigraine agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/14—Drugs for disorders of the nervous system for treating abnormal movements, e.g. chorea, dyskinesia
- A61P25/16—Anti-Parkinson drugs
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/22—Anxiolytics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/24—Antidepressants
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
- A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
Landscapes
- Health & Medical Sciences (AREA)
- Veterinary Medicine (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Biomedical Technology (AREA)
- Neurosurgery (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Engineering & Computer Science (AREA)
- Neurology (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Organic Chemistry (AREA)
- Epidemiology (AREA)
- Pain & Pain Management (AREA)
- Psychiatry (AREA)
- Psychology (AREA)
- Hospice & Palliative Care (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Abstract
Use of neurons, in particular Ventral Tegmental (VTA) Dopamine (DA) capable neurons, anterior Cingulate Cortex (ACC) glutamatergic neurons, gamma-aminobutyric acid (GABA) capable neurons of VTA or dopamine D2 receptors as targets in the preparation of a medicament for the treatment of chronic pain and any of its concomitant symptoms (anxiety, depression, aversion, fear and cognitive impairment).
Description
Technical Field
The invention belongs to the technical field of biological medicine, and particularly relates to application of neurons serving as targets in preparation of products for treating chronic pain and symptoms accompanied with the chronic pain.
Background
Chronic pain is a serious human health hazard, with a short course of disease, for several weeks, up to several years or even throughout the life. Incomplete statistics indicate that about 20-30% of the population worldwide suffers from chronic pain. In addition to a purely painful sensation, chronic pain often causes alterations in mood and cognition such as anxiety, depression, aversion, and the like, and such negative mood and cognitive impairment often enhances pain in the patient. However, the treatment of chronic pain is far behind the clinical need, and there is still a lack of effective analgesic drugs with little side effects. The main common clinical drugs are opium, the drugs treat the symptoms but not the root cause, and the drugs have short curative effect time, quick failure and great side effect. Therefore, searching for the neural mechanisms and therapeutic targets for chronic pain occurrence and long-term maintenance, and aiming at these mechanisms and targets, achieving long-term relief of chronic pain and its negative emotion is a great strategic requirement for national medical health.
The central control mechanism of pain perception in the early stage has been systematically studied, and the control channel and molecular mechanism of the central control mechanism are systematically described. Theory of plastic changes is proposed at various levels from molecular, synaptic, to loop. However, the long-term maintenance of these changes in plasticity requires constant renewal through a process similar to re-consolidation, which is severely dependent on sustained sensory input to the damaged tissue or nerve. Thus, chronic pain remains a puzzle on how it is sustained and aggravated for a long period of time after healing of the injured tissue, and how it is affected by other sensory modalities such as negative emotion and cognitive dysfunction. Thus, the search for new mechanisms and theories for mechanisms of chronic pain is urgent. The principle behind this phenomenon is probably explained by the fact that the positive feedback mechanism is continuously enhanced in the loop after it is activated, which is not clear in the control of chronic pain. This is also the root cause of the non-ideal clinical treatment of chronic pain.
In a large number of clinical treatments, chronic pain patients, in addition to being plagued deeply by pain sensations, are found to be accompanied by a variety of other symptoms such as alterations in mood, sleep disorders and cognition, such as anxiety, depression, aversion, etc. These concomitant symptoms often further enhance the patient's pain sensation, and the pain-concomitant symptoms-vicious circle of pain occurs, further increasing the patient's and social burden. Thus, new targets and related products are urgently needed to treat chronic pain and its concomitant symptoms.
Disclosure of Invention
The invention aims at providing a new target and related products for treating chronic pain and accompanying symptoms thereof.
To achieve the above object, embodiments of the present invention include:
in a first aspect, the invention provides the use of a Ventral Tegmental Area (VTA) dopaminergic neuron (DA-capable neuron), a Anterior Cingulate Cortex (ACC) glutamatergic neuron, a gamma-aminobutyric acid (GABA) capable neuron of the VTA or a dopamine D2 receptor as a target in the manufacture of a medicament for the treatment of any of chronic pain and its concomitant symptoms.
In certain embodiments of the invention, the chronic pain is selected from at least one of trigeminal neuralgia, parkinsonism pain, sciatica, neuropathic pain, migraine and oncologic pain.
In certain embodiments of the invention, the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear, and impairment of cognitive function.
In certain embodiments of the invention, the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
In a second aspect, the present invention provides the use of an enhancer of DA-capable neuronal activity, an inhibitor of ACC glutamatergic neuronal activity or an inhibitor of VTA GABA-capable neuronal activity in the manufacture of a medicament for the treatment of any of chronic pain and its concomitant symptoms.
In certain embodiments of the invention, the chronic pain is selected from at least one of trigeminal neuralgia, parkinsonism pain, sciatica, neuropathic pain, migraine and oncologic pain.
In certain embodiments of the invention, the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear, and impairment of cognitive function.
In certain embodiments of the invention, the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
In a third aspect, the present invention provides the use of a D2 receptor agonist for inhibiting the activity of an ACC glutamatergic neuron projected by a dopaminergic neuron in the receiving Ventral Tegmental Area (VTA), preferably the D2 receptor agonist is selected from at least one of bromocriptine (bromocriptine), a compound of formula (I) and salts thereof, cabergoline (cabergoline), pergolide (pergolide), aspoxicam (apu), pramipexole (pramipexole) and Luo Paini ro (roperlo).
Wherein R1 is a C1-C6 alkyl group, preferably a C1-C3 alkyl group.
In certain embodiments of the invention, the D2 receptor agonist is an inorganic acid salt, preferably a hydrochloride salt, of a compound of formula (I), more preferably the D2 receptor agonist is Quinpirole (Quinpirole) or a salt thereof.
In certain embodiments of the invention, the product is a medicament for treating any of chronic pain and its concomitant symptoms.
In certain embodiments of the invention, the chronic pain is selected from at least one of trigeminal neuralgia, parkinsonism pain, sciatica, neuropathic pain, migraine and oncologic pain.
In certain embodiments of the invention, the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear, and impairment of cognitive function.
In certain embodiments of the invention, the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
In a fourth aspect, the present invention provides the use of a D2 receptor antagonist in the manufacture of a product for acting on a VTA to relieve the self-inhibitory effect of D2 receptors on dopaminergic neurons and to enhance the activity of dopaminergic neurons; preferably, the D2 receptor antagonist is at least one selected from the group consisting of a compound of formula (II) and salts thereof, sulpiride, metoclopramide, domperidone, etipride,
Wherein R1 is a C1-C6 alkyl group, preferably a C1-C3 alkyl group;
r2 and R3 are each independently optionally substituted C6-C12 aryl, preferably halogen substituted C6-C12 aryl, more preferably halogen substituted phenyl, further preferably halogen para-substituted phenyl, further preferably halogen is fluorine or chlorine.
In certain embodiments of the invention, the D2 receptor antagonist is haloperidol (haloperidol) or a salt thereof.
In certain embodiments of the invention, the product is a medicament for treating any of chronic pain and its concomitant symptoms.
In certain embodiments of the invention, the chronic pain is selected from at least one of trigeminal neuralgia, parkinsonism pain, sciatica, neuropathic pain, migraine and oncologic pain.
In certain embodiments of the invention, the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear, and impairment of cognitive function.
In a fifth aspect, the invention provides a pharmaceutical composition comprising a D2 receptor agonist and a D2 receptor antagonist.
In a sixth aspect, the invention provides the use of a pharmaceutical composition of the invention in the manufacture of a medicament for the treatment of any of chronic pain and its concomitant symptoms.
In a seventh aspect, the present invention provides a kit comprising a pharmaceutical composition of the present invention, preferably the kit comprises instructions for administering a D2 receptor agonist and a D2 receptor antagonist for the treatment of any of chronic pain and its concomitant symptoms.
The quinpirole has the following structure:
haloperidol has the following structure:
the inventor makes a major breakthrough in the research and exploration of the mechanism of chronic pain long-term maintenance and other accompanied symptoms. The inventors found that there is a neuro-closed structure between the nociceptive regulatory center anterior gyrus cortex (ACC) and the affective regulatory center Ventral Tegmental Area (VTA) that is participated by 3 kinds of neurons, namely, ACC glutamatergic neurons, gabaergic neurons of the VTA and dopamine neurons of the VTA (DA neurons). And this closed loop structure can implement a positive feedback closed loop between ACC-VTAs through a "double-brake" mechanism. The positive feedback closed loop is not only a continuous and progressive loop mechanism of chronic pain, but also a core mechanism for mediating the generation of associated symptoms of chronic pain. In this positive feedback loop, a progressive decrease in dopaminergic neuronal excitability in the VTA brain region and an increase in glutamatergic neuronal excitability in the ACC brain region trigger the onset of the entire nerve loop, resulting in chronic pain-related concomitant symptoms.
The invention has the following beneficial technical effects:
based on the novel mechanism discovered by the aforementioned inventors, drugs prepared with DA-capable neurons, ACC glutamatergic neurons, gabaergic neurons of VTA, or dopamine D2 receptors as targets can treat any of chronic pain and its concomitant symptoms.
Based on the novel mechanism discovered by the aforementioned inventors, a drug prepared by a DA-capable neuron activity enhancer, an ACC glutamatergic neuron activity inhibitor, or a VTA GABA-capable neuron activity inhibitor can treat any of chronic pain and its concomitant symptoms.
Based on the new mechanism discovered by the aforementioned inventors, products prepared by D2 receptor agonists can be used to inhibit ACC glutamatergic neuronal activity projected by the dopaminergic neurons of the receiving Ventral Tegmental Area (VTA). When the product is a medicament, the medicament may treat any one of chronic pain and its concomitant symptoms.
Based on the novel mechanism discovered by the inventors, products prepared by D2 receptor antagonists can act on VTA to release the self-inhibitory effect of D2 receptors on dopaminergic neurons to enhance dopaminergic neuron activity. When the product is a medicament, the medicament may treat any one of chronic pain and its concomitant symptoms.
Based on the novel mechanism discovered by the foregoing inventors, the present invention provides pharmaceutical compositions comprising D2 receptor agonists and D2 receptor antagonists that can treat any of chronic pain and its concomitant symptoms.
When the medicine prepared by the dopamine D2 receptor agonist or antagonist is injection, powder for injection or tablet for injection, the medicine can treat chronic pain and the accompanying symptoms thereof by acting on different brain areas respectively. Drugs prepared from D2 receptor agonists (e.g., quinpirole) act on the ACC brain region to relieve chronic pain and its concomitant symptoms in an animal model of chronic pain by inhibiting ACC glutamatergic neuronal activity projected by dopaminergic neurons in the receiving Ventral Tegmental Area (VTA). Drugs prepared from D2 receptor antagonists (e.g., haloperidol) act on the brain region of the VTA to treat chronic pain and its concomitant symptoms by restoring activity to the dopaminergic neurons of the VTA.
Drawings
In fig. 1, it is shown that nociception abnormality and its accompanying symptoms are induced in SNI model mice, and decrease in dopaminergic neuron activity in VTA region is caused.
Fig. 2 shows that chemically activating dopaminergic neurons in the VTA area is effective in alleviating chronic pain and its various concomitant symptoms.
Fig. 3 shows that light stimulation inhibits the activation of the dopaminergic neurons in the VTA zone by the glutamatergic neurons in the ACC zone effectively alleviating chronic pain and its various concomitant symptoms.
FIG. 4 shows that injection of haloperidol (a D2R antagonist, 500 nM) into the VTA region, releases the self-inhibitory effect of the D2 receptor on dopaminergic neurons, enhances dopaminergic neuron activity and is effective in alleviating chronic pain and its various concomitant symptoms.
Figure 5 shows that sustained topical administration of haloperidol to the VTA zone can chronically alleviate the nociceptive abnormalities and various concomitant symptoms in the SNI of a chronic pain model mouse.
Fig. 6 shows that SNI model mice caused an enhancement of the glutamatergic neuronal activity in the ACC region.
Fig. 7 shows that chemobiostimulation inhibits the glutamatergic neurons in the ACC region that receive projections of the VTA dopaminergic neurons, effectively and consistently alleviating nociceptive abnormalities and multiple concomitant symptoms in SNI model mice.
Fig. 8 shows that photoactivating dopaminergic neurons, thereby inhibiting the ACC region glutamatergic neurons that receive the DA-capable neuron projections of VTA, effectively alleviates nociceptive abnormalities and multiple concomitant symptoms in SNI model mice.
Fig. 9 shows that the ACC region injected with quinpirole (a D2R agonist, 50 nM) was effective in alleviating chronic pain and its various concomitant symptoms by inhibiting ACC glutamatergic neuronal activity that receives VTA dopaminergic neuronal projections.
Fig. 10 shows that sustained topical administration of quinpirole in the ACC region can alleviate the nociceptive abnormalities and various concomitant symptoms in the SNI of chronic pain model mice over a long period of time.
FIG. 11 shows that biostimulation inhibits GABA energy neuron activity in the VTA region, alleviating dyspain and various concomitant symptoms in SNI model mice.
Detailed Description
The present invention will be described in further detail with reference to specific examples, which are illustrative of the present invention and do not limit the scope of the present invention.
Experiments 1-7 and experimental runs 8-14 required by the present invention are provided below.
1. Sciatic nerve branch selective injury model (SNI)
After the mice were anesthetized with isoflurane, the right thigh side skin was incised, and the biceps femoris was incised. 3 branches of the exposed sciatic nerve: sural nerve, common sural nerve and tibial nerve. The common fibular nerve and tibial nerve were ligated with non-absorbable sutures at the trigeminal points, and the nerve was then severed to the distal end of each junction and the distal end was resected 3-5 mm. Suturing the muscles and skin. The Sham group was exposed only to the right sciatic nerve and not ligated. Mice with paralysis, dyskinesia, or death of lower extremities after surgery in the pain model group will be removed from the experiment, replaced with new mouse modeling and added to the group. Pre-and post-operative 14d mechanical and thermal pain sensitivity were assessed. Model mice model side hindpaws were unable to develop normally, whereas sham operated mice did not.
2. Mechanical pain sensitivity (or touch induced pain):
the mechanical foot-constricting reflex threshold (mechanical withdrawal threshold, MWT) of the left and right hind limbs of the mice was measured by Von Frey method, the mice were placed in an plexiglas cage with a metal small lattice net at the bottom. After the animal is in a calm state after adapting to the environment and stopping exploring the behavior, the Von Frey filaments with different gram weights are used for stimulating the midfoot center part of the sole, and the behavior of lifting or licking feet of the mouse is regarded as positive reaction. The heavier the Von Frey filaments, the higher the pain threshold, the less painful the mice; the smaller the Von Frey filaments, the lower the pain threshold, the more painful the mice.
3. Cold and hot plate experiment:
the super thermostat is opened, and the temperature of the super thermostat is regulated to be constant at 0+/-0.1 ℃/50+/-0.1 ℃. The test mice were selected and qualified subjects were pre-selected before the test. Mice were placed one by one in the measuring instrument, and first the normal value was measured. The pain threshold was then measured once each after mice were grouped and recorded. The less the mice stay in the tester, the lower the pain threshold, and the more painful the mice.
4. Pain anxiety emotional behavioral testing:
overhead plus maze experiment: the mice were placed on an elevated plus maze center platform facing one of the open arms. The mice were allowed to freely explore the four arms for 5min, and the latency of the mice to enter any arm for the first time was recorded, with the four limbs of the mice all entering one arm as one entry. The number of times the mice entered the open arm and the residence time in the open arm were calculated as the percentage of the total number of times the mice entered the open arm and the residence time in the closed arm (sum of the number of times the mice entered the open arm) and the total time (sum of the residence time in the open arm and the residence time in the closed arm), respectively, and were used as an index for evaluating anxiety. The fewer times the mice enter the open arm and the less time they stay in the open arm, the more anxious the mice are.
Open field experiments: the mice were placed in a square box (30X 30 cm) with transparent plexiglas at the periphery and bottom and allowed to freely move for 15min, and the total movement distance and movement time of the mice in the experimental box were measured. 15X 15cm in the middle of the experiment box 2 The region is the central region and occupies one quarter of the total area. Each bin only allowed one mouse to be tested at a time, with the time of mice activity in the central zone as an indicator of evaluation anxiety. The less time the mice were active in the central zone, the more anxious.
5. Pain depression emotional behavioural detection:
sucrose preference experiment: mice were kept individually. Prior to testing, mice were habituated to two bottles of the same 1% sucrose for 2 days, followed by 2 days with water, respectively. After acclimation, the mice were deprived of moisture for 24 hours. The mice were then free to drink either 1% sucrose solution or plain boiled water for 24 hours. To avoid positional preference, the bottle positions were exchanged half the time during the test. Sucrose preference is calculated as a percentage of sucrose consumption to the total consumption of liquid. The less sucrose consumption, the more depressed the mouse state.
Forced swimming experiment: a transparent cylinder (20 cm in height, 15cm in diameter) was used to fill 23-25℃water to a depth of 15 cm. The behavior of the mice was recorded from the side with a camera for 6 minutes. Analysis was performed for the last 5 minutes and the mice were considered to be inactive when they were floating or kept stationary. The longer the immobility time of the mice, the more depressed.
6. Aversion emotion behavioural detection:
conditional locality preference (CPP) experiments were used for aversion emotional behavior detection. The test was performed using a three-chamber box. The wall of the left room is characterized by vertical black and white stripes, and the floor is a strip-shaped perforation. The middle room is an ash wall and is a short strip-shaped punched floor slab. The right room is a black wall and a round perforated floor. CPP duration was 5 days. The pre-test was performed on the first day. In the pre-test, the mice entered all three rooms for 15 minutes and the time of the mice in each room was recorded. Day 2-4, CPP test, mice were allowed to move in the right/left ventricle for 40 minutes immediately after intracranial injection of CNO, and after 4 hours, mice were allowed to move in the contralateral ventricle for 40 minutes immediately after intracranial injection of normal saline. Post-conditioning experiments were performed on day 5 without CNO/saline injection. The mice can enter all three rooms and record the time they stay in each room. Movement of mice was recorded with SMART software. CPP score was calculated from the time of mice in right/left chamber (CNO side) versus the time of mice in left/right chamber (saline side). The longer the residence time of the mice in the CNO injection chamber, the stronger the preference of the mice.
7. Learning cognitive behavioral testing:
Y maze: animals were placed in the starting arm of the Y maze and allowed to explore freely. At this stage, the researcher may record the animal's behavior, including its residence time in the various arms, steering options, and the like. The higher the ratio of mice selected between the arms, the more learning and cognitive ability the mice were shown to be.
New object identification: a mouse was placed in a cage with two identical objects for 10 minutes. The mice were then returned to their reared cages for one hour. Then after a sample object is replaced with a new (novel) object, it is returned to the cage. It is determined how much time the animal has interacted with the new object. The longer the new object contact time, the stronger the learning and cognitive ability of the mice.
8. In vivo and ex vivo optogenetic and chemogenetic
For in vivo optogenetic experiments, the inventors allowed mice to express virus for the first week after injection of virus in the corresponding brain region, buried the fiber in the VTA/ACC region, and recovered the mice for one week and five minutes per day of palpation and adaptation, which operation was continued for three days. After the fiber and the head ferrule are docked, the mice are placed in a behavioural detection device, and a laser is used to activate the already integrated light sensitive protein and detect behaviours. The lasers are preferably blue (488 nm laser) and yellow (596 nm laser) lasers from the family Qian-Oxing, with lasers having a wavelength of 488nm/596 nm. For ex vivo optogenetics, the inventors also used a laser with a wavelength of 488nm/596nm in the corresponding brain region to activate the already integrated light sensitive protein and record the neuronal response. Prior to the experiment, the inventors used a power meter (Thorlabs) to adjust the laser intensity. In this experiment, the laser intensity of the inventors was about 10-15 milliwatts (mW).
For in vivo chemistry genetics experiments, the experimental procedure was essentially identical to optogenetics, except that a cannula was embedded in the VTA/ACC region, and the mice were given Clozapine (CNO) (e.g. C0832 from Sigma) via the cannula and tested for behaviours after one week recovery. For the experiments of ex vivo chemistry genetics, the inventors prepared ex vivo brain tablets after injecting the virus in the corresponding brain region, after proper expression of the mouse virus, administered CNO in the VTA region, and recorded the response to activating the nerve.
9. Carbon fiber electrochemical recording
Electrochemical recordings were made using 200 μm long, 7 μm diameter micro Carbon Fiber Electrodes (CFEs). The carbon fiber electrode was inserted with its exposed tip fully into the ACC brain sheet at a 30 ° tilt angle, 300 μm from the stimulating electrode. The voltage of the recording electrode was stabilized at 780mV using an EPC9/2 amplifier and Pulse software (HEKA Electronic, lambrecht/Pfalz, germany). The stimulation electrode was a bipolar platinum wire electrode (diameter 150 μm, plastics One Inc., USA) produced by a Master-8 stimulator (AMPI, israel). Each pulse was 0.2ms in duration, 0.6mA in amplitude, and 3min per time interval of two stimulations. The collection frequency of the ampere signal (Iamp) was 3.13kHz and was low frequency filtered using 100Hz and the Off-line analysis was performed using Igor software (WaveMetrix).
10. Immunohistochemistry
After the animal is anesthetized, the animal is perfused by physiological saline and 4% paraformaldehyde heart respectively, the brain is taken out and soaked for the same 4% polymer for 6 hours, then sugar is deposited for 3 days (4 ℃), 30 mu m thick slices are cut from each tissue, conventional bleaching and dyeing are carried out by a two-step method, the tissue slices are incubated by corresponding acceptor antibody serum respectively, the second antibody is used for overnight, a laser confocal microscope is used for imaging, and the immunohistochemical dyeing result is observed.
11. Whole-cell patch clamp
Brain pieces incubated for 1 hour are moved into a recording tank, mixed gas saturated ACSF (patch clamp special) is circulated and perfused, the temperature is maintained at 28-30 ℃, and the flow rate is 1.5ml/min. And (3) observing under a forward infrared microscope (BX 51W 1), and selecting and recording the dopaminergic neurons, GABAergic neurons and glutamatergic neurons in the VTA area. Spontaneous and evoked Action Potentials (APs) were recorded in current clamp mode using a 200B amplifier. The series resistance was monitored throughout the experiment, only those data with series resistance <30mΩ and fluctuation less than 20% were available for analysis. All recordings were performed under the Axon 200B patch clamp amplifier, digital to analog converter Digidata 1440A, stimulator Grass, and pCLAMP 10.3 software system with a sampling frequency of 10000Hz. The mEPSCs data were analysed using mini Analysis software.
12. Viral injection or burial
After anesthetizing the mice with 3% isoflurane, the cranium tops were dehaired, fixed on a stereotactic injector, the cranium top skin was cut open, the cross-stitch was exposed, three-dimensional parameters were adjusted, holes were drilled with the small animal cranium, and the virus was injected at the injection site using injection devices known in the art. Preferably, the injection device is a microinjector, through which 0.5ml of virus is injected at a rate of 0.1ml/min per site, and left for 5min after the injection is completed, and the microinjector is removed. And the buried pipe is used for feeding, a stainless steel pipe (with the outer diameter of 0.5 mm) is buried in the positioning area, the probe is fixed by using dental tray powder, the incision is closed, and the animal is placed in a cage for normal feeding after waking up. After 3% isoflurane anesthesia of mice prior to behavioural testing, physiological saline or the corresponding agonists and inhibitors were administered separately using a microinjection pump, and behavioural testing was performed.
13. Preparation of the injection:
haloperidol (company: sigma cat# H1512) powder was dissolved in dimethyl sulfoxide (DMSO) or glycerol to prepare 500. Mu.M stock solution, and the test sample was prepared into an injection in a drug delivery vehicle (artificial cerebrospinal fluid (ACSF)) at a concentration of 500nM for electrophysiological experiments. Haloperidol powder was dissolved in Dimethylsulfoxide (DMSO) or glycerol to prepare 500 μm stock solution, and the test sample was prepared into injections in a drug delivery vehicle (physiological saline) at a concentration of 50 μm for use in the behavioury of pain and associated concomitant symptoms. A50. Mu.M stock solution was prepared by dissolving quinpirole (company: sigma cat# Q102) powder in physiological saline, and the test substance was prepared into injections in a drug delivery vehicle (artificial cerebrospinal fluid (ACSF)) at a concentration of 50nM for electrophysiological experiments. The preparation method comprises dissolving quindox Luo Fenmo in physiological saline to obtain 10 μg/μl stock solution, and preparing the test sample into injection in administration carrier (physiological saline) at concentration of 1 μg/μl for pain and related symptomatic behaviours. Clozapine (CNO: sigma cat# C0832) powder was dissolved in physiological saline to prepare 10mg/kg stock solution, and the test sample was prepared into injections at a concentration of 0.5mg/kg in a carrier for administration (physiological saline) for pain and related symptomatic behaviours.
14. Statistical analysis
Data are expressed as mean ± SEM. The data were analyzed using Prism (GraphPad). Two pairs of the two-tailed student T tests are adopted for comparison. Other data correct for multiple comparisons using one-or two-factor analysis of variance. In all cases, p less than 0.05 is considered significant.
The following examples serve to demonstrate that targeting specific neurons and related receptors can treat chronic pain and its concomitant symptoms.
Example one Effect of treating chronic pain and its accompanying symptoms Using dopaminergic neurons of the VTA region as the target
Relationship between VTA dopaminergic neurons and chronic pain and accompanying symptoms in SNI model mice
The inventors constructed a chronic pain model of sciatic nerve branch injury (SNI) on C57 mice (fig. 1), a shows the mechanical and thermal pain response mediated by mice tested by Von Frey model SNI (n=10 vs mice, <0.001, < p). SNI model mice showed significant mechanical nociceptive abnormalities and were able to sustain for up to 6 weeks or more, b showed SNI model-mediated thermal nociceptive responses (n=7 vs mice, ×p < 0.01). The SNI model can well simulate the pain sensitivity abnormality of chronic pain.
The inventors subsequently tested whether chronic pain induced the associated negative emotion in model mice by a related anxiety behavior experiment. Experimental results showed that c-f are SNI model mice that showed significant anxiety-like behavior in both open field and overhead experiments (n=6 pairs of mice, two groups with p <0.01 and p < 0.05).
To investigate whether altered function of dopaminergic neurons in the VTA zone plays a key role in pain development, the inventors examined whether the synaptic transmission function of SNI model mice was altered by electrophysiological-whole cell patch clamp technique, and g results showed that the frequency of model mice action potential was significantly reduced, indicating that their cell activity was significantly reduced (n=14 cells from 7 pairs of mice, in two groups, p < 0.01). The results demonstrate that dopaminergic neurons in the VTA zone play an important regulatory role in the development and progression of chronic pain (fig. 1).
The results of the a-g correlation experiments in FIG. 1 (all expressed as the mean value of each group) are shown below:
2. chemical stimulation to control activity of dopaminergic neurons in VTA region for relieving chronic pain and accompanying symptoms
To verify that manipulation of dopaminergic neuron activity in the VTA zone is an important target for the regulation of chronic pain and its concomitant symptoms, the inventors validated by virus injection, electrophysiology and behavioural (fig. 2). The results show that the chemogenetic activation virus injected in the VTA region is activated by intraperitoneal injection of Clozapine (CNO) so as to specifically activate the activity of dopaminergic neurons in the VTA region, and then the chemogenetic activation virus can effectively relieve chronic pain and various accompanying symptoms such as dysphoria, depression, aversion and awareness (figure 2).
A-b in fig. 2 shows that chemogenetically activated virus (pAAV-EF 1 a-DIO-hM 3D (Gq) -EGFP-WPRE company: OBIO cat No. H15959) injected in the VTA zone, which enhances dopaminergic neuron action potential release by intraperitoneal injection of Clozapine (CNO) drug activated virus (n=15 cells from 8 pairs of mice, two groups, p <0.01,); chemically activating the virus to activate dopaminergic neuron activity in the VTA zone is effective to alleviate mechanical and thermal nociceptive responses in SNI mice (n=8 vs mice, <0.05, <0.01, <0.001, < p); e-i is that chemogenetically activated virus activation of dopaminergic neuronal activity in the VTA zone is effective to alleviate pain associated with aversion-like emotional response, anxiety-like emotional response, depression-like emotional response and cognitive impairment behavior in SNI mice (n=10 vs mice, two groups, p <0.05, p <0.01, p < 0.001).
The results of the a-h correlation experiments in FIG. 2 (all expressed as the mean value of each group) are shown below:
/>
/>
these experimental results further demonstrate that abnormalities in dopaminergic neuron activity in the VTA zone play an important role in the development and progression of chronic pain. And the regulation and control of the activity of the compound can effectively relieve the occurrence of dyspain and various accompanying symptoms of chronic pain. The dopaminergic neurons of the VTA zone can be used as targets in the preparation of a medicament for the treatment of any of chronic pain and its concomitant symptoms. The medicament prepared from the DA energy neuron activity enhancer can treat any one of chronic pain and accompanying symptoms thereof.
3. Electrical stimulation control of dopaminergic neuron activity in VTA region for relieving chronic pain and accompanying symptoms
To further verify that the electrical stimulation pattern can also regulate the occurrence and development of chronic pain and its concomitant symptoms by manipulating dopaminergic neuron activity in the VTA zone, the inventors verified by virus injection, electrophysiology and behavioural (fig. 3). The results show that the dopaminergic neuron in the VTA region specifically inhibits the activity of the glutamatergic neuron in the ACC region through optogenetics inhibiting virus and optical stimulation and reduces the inhibiting effect on the dopaminergic neuron in the VTA region, and the activity of the dopaminergic neuron in the DA region is activated, so that the dopaminergic neuron can effectively relieve various accompanying symptoms such as dysphoria, anxiety, depression, aversion, cognition and the like of chronic pain (figure 3).
A-c in fig. 3 shows that optogenetically-inhibited virus (pAAV-EF 1 a-DIO-eNpHR 3.0-mCherry-WPRE company: OBIO cat No. H4882) is injected into ACC region, and yellow light (596 nm) of laser transmits light stimulus to ACC brain region through optical fiber embedded in mouse ACC brain region, thereby inhibiting glutamate neuron activity in ACC region and weakening inhibitory effect on dopamine neuron in VTA region, thereby enhancing dopaminergic neuron activity (glutamate neuron in ACC region projects and activates gabaergic neuron in VTA region, gabaergic neuron inhibits dopaminergic neuron activity in VTA region), and effectively relieving mechanical and thermal sensitization response of SNI mice (n=8 vs. mice, two groups, p <0.01, p < 0.001); d-h is the optogenetically inhibiting projection of the glutamatergic neurons of the ACC region to the VTA region, thereby enhancing the dopaminergic neuron activity of the VTA region and effectively alleviating pain associated with anxiety-like, aversive, depression-like and cognitive impairment behaviors in SNI mice (n=10 vs. mice, two groups, p <0.05, p <0.01, p < 0.001).
The results of the a-g correlation experiments in FIG. 3 (all expressed as the mean value of each group) are shown below:
/>
the experimental results further show that the electrical stimulation regulates and controls the activity of dopaminergic neurons in the VTA area, and can effectively relieve the pain-sensitivity abnormality of chronic pain and the occurrence of various accompanying symptoms. The dopaminergic neurons of the VTA zone can be used as targets in the preparation of a medicament for the treatment of any of chronic pain and its concomitant symptoms. The medicament prepared from the DA energy neuron activity enhancer can treat any one of chronic pain and accompanying symptoms thereof.
Evaluation of the effects of d2 receptor antagonists on the treatment of chronic pain and various concomitant symptoms
(1) Evaluation of the effects of D2 receptor antagonists on dopaminergic neurons in the VTA region
The D2R antagonist haloperidol and 8 pairs of 6-8 week old model mice prepared by experiment 1 were used for this evaluation.
Haloperidol powder was dissolved in Dimethylsulfoxide (DMSO) or glycerol to prepare 500 μm stock solution, and the test sample was prepared into an injection in a drug delivery vehicle (artificial cerebrospinal fluid (ACSF)) at a concentration of 500 nM.
Separate brain slices were prepared by head-cutting the model mice and the test mice (8 mice each), brain slices containing VTA were selected, action potentials were recorded after clamping dopaminergic neurons in the VTA zone according to the above-described procedure 11, and the above-described injections (test group)/artificial cerebrospinal fluid (model group) were added to the brain slices through an electrophysiological administration system. According to experiment 11, action potentials were recorded and the dopaminergic neuron activity in the VTA region was determined.
A-b in fig. 4 shows that local injection of haloperidol (a D2R antagonist) in the VTA region increases the dopaminergic neuron AP firing frequency in the VTA region (haloperidol: 500nm, n=10 cells from 8 mice, <0.001,/p)
The results of the correlation experiments (all expressed as means of each group) are shown below:
(2) Evaluation of the effect of D2 receptor antagonists on chronic pain
The D2R antagonist haloperidol and 8 pairs of 6-8 week old model mice prepared by experiment 1 were used for this evaluation. Mice were divided into 2 groups (model group and test group), 8 each.
The haloperidol powder was dissolved in dimethyl sulfoxide (DMSO) or glycerol to prepare a 500. Mu.M stock solution, and the test sample was prepared into an injection in a carrier (physiological saline) at a concentration of 50. Mu.M.
According to the foregoing operation 12, saline was locally injected in the VTA area by a microinjector for the model group, and the foregoing injection (50 μm,200 nl) was locally injected in the VTA area by a microinjector for the test group. After injection of the aforementioned injections for 3 minutes, the mechanical nociceptive response was determined according to experiment 2.
The experimental results demonstrate that the mechanical nociceptive response (haloperidol: 50 μm,200 nL) of SNI mice is relieved (n=8 vs. mice, two groups, p < 0.01).
The results of the correlation experiments (all expressed as means of each group) are shown below:
(3) Evaluation of the Effect of D2 receptor antagonists on treating various concomitant symptoms of chronic pain
The D2R antagonist haloperidol and 20 model mice prepared from experiment 1, 6-8 weeks old, were used for this evaluation. Mice were divided into 2 groups (model group and test group), 10 each.
The haloperidol powder was dissolved in dimethyl sulfoxide (DMSO) or glycerol to prepare a 500. Mu.M stock solution, and the test sample was prepared into an injection in a carrier (physiological saline) at a concentration of 50. Mu.M.
According to the foregoing operation 12, saline was locally injected in the VTA area by a microinjector for the model group, and the foregoing injection (50 μm,200 nl) was locally injected in the VTA area by a microinjector for the test group. After 3 minutes of injection, anxiety-like behavior, depression-like behavior, aversion-like behavior and cognitive behavior of mice were determined according to experiments 4-7.
The results of the correlation experiments (all expressed as means of each group) are shown below:
/>
d-h of fig. 4 shows that SNI mice were effectively relieved of anxiety-like behavior, depression-like behavior, aversion-like behavior, and cognitive behavior following topical administration of haloperidol to the VTA (n=10 vs mice, two groups, p <0.05, p < 0.01). Thus, haloperidol has been shown to treat a variety of concomitant symptoms of chronic pain.
(4) Evaluation of long-term effects of D2 receptor antagonists on treatment of multiple concomitant symptoms of chronic pain
The D2R antagonist haloperidol and 10 pairs of model mice prepared from experiment 1 6-8 weeks old were used for this evaluation. Mice were divided into 2 groups (model group and test group), 10 each.
The haloperidol powder was dissolved in dimethyl sulfoxide (DMSO) or glycerol to prepare a 500. Mu.M stock solution, and the test sample was prepared into an injection in a carrier (physiological saline) at a concentration of 50. Mu.M.
According to the foregoing operation 12, saline was locally injected in the VTA area by a microinjector for the model group, and the foregoing injection (50 μm,200 nl) was locally injected in the VTA area by a microinjector for the test group. Mice were assayed for nociceptive and anxiety-like behavior according to experiments 2 and 5 after 2 weeks of continuous haloperidol injection (once every other day of injection) in the VTA. A in fig. 5 is a schematic representation of repeated application of haloperidol in the VTA zone for 2 weeks (once every other day of injection);
the model mice and the test mice (7 mice each) were separately subjected to head-breaking to prepare ex-vivo brain slices, brain slices containing VTA were selected, action potentials were recorded after clamping dopaminergic neurons in the VTA area according to the above-described procedure 11, and the above-described injections (test group)/artificial cerebrospinal fluid (model group) were added to the brain slices through an electrophysiological administration system. According to experiment 11, action potentials were recorded and the dopaminergic neuron activity in the VTA region was determined.
The results of the correlation experiments (all expressed as means of each group) are shown below:
/>
figure 5 b shows pain-sensitive relief from SNI with haloperidol applied for 2 weeks (once every other day of injection) and for at least 1 week after stopping haloperidol application (n=8 vs mice, two groups, p < 0.001).
Figure 5 c shows that the action potential firing rate of haloperidol on dopaminergic neurons in the VTA zone is significantly reduced (n=14/7 vs. mice, two groups, p < 0.05).
D-e of fig. 5 shows that anxiety-like behavior of SNI mice was effectively alleviated after haloperidol application (n=8 mice in two groups with p <0.05, p < 0.01);
these results (fig. 5) demonstrate that haloperidol antagonizes the self-inhibitory effect of dopaminergic neurons D2R in the VTA zone on dopaminergic neurons, thereby enhancing dopaminergic neuron activation, is effective in alleviating pain, and has a durable efficacy against pain and its concomitant symptoms, particularly for at least 1 week after stopping haloperidol injection.
The above experimental results demonstrate that: the dopaminergic neurons of the VTA zone can be used as targets in the preparation of a medicament for the treatment of any of chronic pain and its concomitant symptoms. The medicament prepared from the DA energy neuron activity enhancer can treat any one of chronic pain and accompanying symptoms thereof.
Example two, treatment of chronic pain and its concomitant symptoms with ACC glutamatergic neurons as target
1. SNI model mice elicit enhanced ACC glutamatergic neuronal activity
To investigate whether functional changes in glutamatergic neurons in the ACC region play a key role in the development of pain, the inventors examined whether the synaptic transmission function of SNI model mice was altered by electrophysiological-whole cell patch clamp technique, and the results in fig. 6d showed that the frequency of model mice action potentials was significantly increased, indicating that their cellular activities were significantly decreased (n=16 cells from 7 pairs of mice, in two groups, p < 0.01). The results demonstrate that the glutamatergic neurons of the ACC region play an important regulatory role in the development and progression of chronic pain (fig. 6).
A-b in fig. 6 are SNI model mice mediated mechanical and thermal pain responses (n=10 mice, p <0.01, p < 0.001); c is that a reverse standard virus (rAAV-hSyn-CRE-mCherry-WPRE-hGH pA company: brainVTA product number: PT-0407) is injected into the VTA area, and the slice staining-free result of the ACC area is shown schematically, which shows that glutamatergic neurons of the ACC area are projected to the VTA area; d shows an increased frequency of glutamatergic neural action potential firing in the ACC region of SNI mice, i.e. an increased glutamatergic neural activity (n=16 cells/7 pairs of mice, two groups, p < 0.01).
The results of the correlation experiments (all expressed as means of each group) are shown below:
/>
the results of the inventors (fig. 6) show that ACC glutamatergic neuronal excitability is significantly enhanced in SNI model mice. And the research of the inventor discovers that the inhibition effect of the reduction of the activity of the dopaminergic neurons on the glutamatergic neurons of the ACC in the SNI model mouse is obviously reduced for the first time, so that the activity of the glutamatergic neurons is excessively enhanced.
2. Chemical stimulation to control ACC glutamatergic neuron activity to relieve chronic pain and its accompanying symptoms
The study of the inventor finds that a nerve closed loop structure participated by 3 kinds of neurons (ACC glutamatergic neuron, VTA GABA (gamma-associated GABA) and VTA dopaminergic neuron) exists between a control center Anterior Cingulate Cortex (ACC) of pain sense and a feeling control center Ventral Tegmental Area (VTA). Glutamatergic neurons of the ACC region indirectly regulate dopaminergic neurons of the VTA region through the VTA gabaergic neurons, which in turn project back into the originating ACC glutamatergic neurons.
Therefore, when the inventor activates the virus and Clozapine (CNO) drug specifically through chemogenetics to activate VTA dopaminergic neuron, thereby inhibiting the activity of ACC region glutamatergic nerve, it can effectively relieve chronic pain and various accompanying symptoms such as anxiety, depression, aversion and awareness (figure 7).
In fig. 7, a-b show that injection of chemogenetically activated virus (pAAV-EF 1 a-DIO-hM 3D (Gqh) -EGFP-WPRE company: OBIO cat No. H15959) in the VTA zone enhances dopaminergic neuron activity by injecting Clozapine (CNO) drug in the ACC brain zone, resulting in reduced excitability of glutamatergic neurons projected to the ACC zone to receive modulation of dopaminergic neurons in the VTA zone, thereby effectively alleviating mechanical and thermal nociceptive responses in SNI mice (n=8 vs, p <0.01, p < 0.001); c-g shows that chemogenetically activated viruses inhibit the decrease in the frequency of release of the action potential of the glutamatergic neurons of ACC, and the decrease in the activity of the neurons, and can effectively relieve the pain associated with aversion-like emotional response, anxiety-like emotional response, depression-like emotional response and cognitive impairment behavior in SNI mice (n=10 vs. mice, in two groups, p <0.05, p < 0.01).
The results of the a-g correlation experiments in FIG. 7 (all expressed as the mean value of each group) are shown below:
/>
these experimental results further demonstrate that abnormalities in ACC glutamatergic neuronal activity play an important role in the development and progression of chronic pain. And the regulation and control of the activity of the compound can effectively relieve the occurrence of dyspain and various accompanying symptoms of chronic pain.
3. Electric stimulation control of ACC glutamatergic neuron activity for relieving chronic pain and accompanying symptoms
To further verify that the electrical stimulation pattern can also regulate the occurrence and development of chronic pain and its concomitant symptoms by manipulating ACC glutamatergic neuronal activity, the inventors verified by viral injection, electrophysiology and behavioural (fig. 8). The results showed that, after specific activation of dopamine nerves in VTA region and projection to ACC by optogenetic activation of viruses and photostimulation and inhibition of ACC glutamatergic neuron activity, it was effective in alleviating pain-sensitive abnormalities of chronic pain and various accompanying symptoms such as anxiety, depression, aversion and awareness (fig. 8).
A-b in fig. 8 shows that injection of labeled cis-labeled virus (pAOV-CAG-EGFP-2A-CRE company: OBIO cargo number: CN 396) in ACC to label glutamatergic neurons of ACC projected onto VTA zone, injection of labeled virus and optogenetically activated virus (pAAV-EF 1a-DIO-WGA-NLS-Flpo-WPRE company: OBIO cargo number: H18411; rAAV-TH-fDIO-hChR2 (H134R) -mCherry-WPRE-hGH p company: brainVTA cargo number: PT-2594) in ACC buried optical fiber, transmission of optical stimulus to ACC brain zone by blue light (488 nm) of laser through optical fiber buried in mouse ACC brain zone, activation of virus by optical stimulus can enhance dopaminergic neuron activity and its projection onto ACC, whereas dopaminergic neuron activity enhancement effectively inhibits glutamatergic neuron action potential release frequency, inhibits activity and can alleviate both i and i effects on mouse pain (sna-16.01; 1.0.01 to two mice, =0.01,); c-g shows that optogenetically activating dopaminergic neurons in the VTA zone and inhibiting ACC glutamatergic neuronal activity is effective in alleviating pain associated with anxiety-like emotional responses, aversion-like emotional responses, depression-like emotional responses, and cognitive impairment behavior in SNI mice (n=10 vs mice, two groups, p <0.05, p <0.01, p < 0.001).
The experimental results further show that the ACC glutamatergic neuron can effectively regulate and control the activity of the ACC glutamatergic neuron, and can effectively relieve the abnormal pain sensitivity of chronic pain and various accompanying symptoms.
The results of the a-g correlation experiments in FIG. 8 (all expressed as the mean value of each group) are shown below:
/>
/>
3. evaluation of the efficacy of postsynaptic D2 receptor agonists in the ACC region for the treatment of chronic pain and various concomitant symptoms
(1) Evaluation of effects of D2 receptor agonists on glutamatergic neurons
7 pairs of model mice prepared from experiment 1, 6-8 weeks old, were used for this evaluation.
A50. Mu.M stock solution was prepared by dissolving quinpirole (company: sigma cat# Q102) powder in physiological saline, and the test substance was prepared into injections in a drug delivery vehicle (artificial cerebrospinal fluid (ACSF)) at a concentration of 50nM for electrophysiological experiments.
Separate brains were prepared by breaking the heads of model mice and test mice (7 mice each), and the brains containing ACC were selected and, according to the procedure 11 described above, after clamping glutamatergic neurons in the ACC region, action potentials were recorded, and the injections (test group)/artificial cerebrospinal fluid (model group) described above were added to the brains by an electrophysiological administration system. According to experiment 11, action potentials were recorded and the glutamatergic neuronal activity in the ACC region was determined.
The results of the correlation experiments (all expressed as means of each group) are shown below:
a-b in fig. 9 shows a decrease in the frequency of glutamatergic neuronal action potential discharge in ACC brain tablets with topical administration of quinpirole (a D2R agonist, 50 nM) (n=10 cells from 7 pairs of mice, < p < 0.001).
(2) Evaluation of the effect of D2 receptor agonists on chronic pain
14 model mice prepared from experiment 1, 6-8 weeks old, were used for this evaluation. Mice were divided into 2 groups (model group and test group), 7 each.
The preparation method comprises dissolving quindox Luo Fenmo in physiological saline to obtain 10 μg/μl stock solution, and preparing the test sample into injection in administration carrier (physiological saline) at concentration of 1 μg/μl for use in pain-sensitive behavioural.
According to the foregoing operation 12, saline was locally injected in the ACC region by a microinjector for the model group, and the foregoing injection (1 μg/μl,200 nl) was locally injected in the ACC region by a microinjector for the test group. After injection of the aforementioned injections in ACC for 3 minutes, the mechanical nociceptive response was determined according to experiment 2.
The results of the correlation experiments (all expressed as means of each group) are shown below:
agonists were demonstrated to be effective in alleviating nociceptive abnormalities in SNI model mice (quinpirole: 1 μg/μl,200 nl) (n=7 vs, two groups of mice, < 0.001). Local injection of the D2R agonist quinpirole in ACC alleviates mechanical pain in SNI mice.
(3) Evaluation of the effect of D2 receptor agonists on the treatment of various concomitant symptoms of chronic pain
20 model mice prepared from experiment 1, 6-8 weeks old, were used for this evaluation. Mice were divided into groups (model and test groups) of 10 mice each.
The preparation method comprises dissolving quindox Luo Fenmo in physiological saline to prepare 10 μg/μl stock solution, and preparing the test sample into injection in administration carrier (physiological saline) at concentration of 1 μg/μl for use in behavioural of pain-associated concomitant symptoms.
According to the foregoing operation 12, saline was locally injected in the ACC region by a microinjector for the model group, and the foregoing injection (1 μg/μl,200 nl) was locally injected in the ACC region by a microinjector for the test group. After injection of the aforementioned injections in ACC for 3 minutes, anxiety-like behavior, depression-like behavior, aversion-like behavior and cognitive behavior of mice were determined according to experiments 4-7.
The results of the correlation experiments (all expressed as means of each group) are shown below:
/>
/>
d-h of fig. 9 shows that SNI mice were effective in alleviating anxiety-like, depression-like, aversion-like and cognitive behaviors following topical administration of flupirome to ACC (n=10 vs. mice, in two groups, p <0.05, p <0.01, p < 0.001). Local injection of the D2R agonist quinpirole in ACC was demonstrated to alleviate anxiety-like, depression-like, aversion-like and cognitive behaviors (fig. 9).
(4) Evaluation of long-term effects of D2 receptor agonists on chronic pain and its various concomitant symptoms
16 model mice prepared from experiment 1, 6-8 weeks old, were used for this evaluation. Mice were divided into groups (model and test groups) of 8 mice each.
The preparation method comprises dissolving quindox Luo Fenmo in physiological saline to obtain 10 μg/μl stock solution, and preparing the test sample into injection in administration carrier (physiological saline) at concentration of 1 μg/μl for pain and related symptomatic behaviours.
According to the foregoing operation 12, saline was locally injected in the ACC region by a microinjector for the model group, and the foregoing injection (1 μg/μl,200 nl) was locally injected in the ACC region by a microinjector for the test group. Mice were assayed for nociceptive and anxiety-like behavior according to experiments 2 and 5 after 2 weeks of continuous injection of quepirole in ACC (once every other day). A in fig. 10 is a schematic diagram of repeated application of quinpirole in the ACC region for 2 weeks (once every other day of injection).
The results of the correlation experiments (all expressed as means of each group) are shown below:
fig. 10 b shows pain-sensitive relief from SNI with 2 weeks (every other day) of quinpirole application, and for at least 1 week of relief after cessation of quinpirole application (n=8 mice, <0.001, < p); fig. 10 c-d show that anxiety-like behavior of SNI mice was effectively alleviated after application of quinpirole (n=8 mice, <0.05, <0.01, < p);
The continuous pain caused by SNI and its concomitant symptomatic behavior showed progressive efficacy after repeated application of quinpirole in ACC for 2 weeks and continued relief for at least 1 week after stopping haloperidol application (fig. 10).
These results indicate that activation of D2R on ACC glutamatergic neurons is effective in alleviating pain by inhibiting ACC glutamatergic neuron activity and has a durable therapeutic effect on persistent pain and co-morbid multiple concomitant disorders. The above experiments define postsynaptic D2R in ACC as a therapeutic target for the treatment of persistent pain and co-morbid end disorders.
The above experimental results demonstrate that: the anterior cingulate cortex glutamatergic neuron can be used as a target for preparing medicines for treating any one of chronic pain and accompanying symptoms. Medicaments prepared from inhibitors of ACC glutamatergic neuronal activity can treat any of chronic pain and its concomitant symptoms.
Example III effects of treating Chronic pain and its concomitant symptoms Using VTA GABAergic neurons as targets
The study of the inventor finds that a nerve closed loop structure participated by 3 kinds of neurons (ACC glutamatergic neuron, VTA GABA (gamma-associated GABA) and VTA dopaminergic neuron) exists between a control center Anterior Cingulate Cortex (ACC) of pain sense and a feeling control center Ventral Tegmental Area (VTA). Glutamatergic neurons of the ACC region indirectly regulate dopaminergic neurons of the VTA region through the VTA gabaergic neurons, which in turn project back into the originating ACC glutamatergic neurons. The inventors have previously found that the activity of class 2 neurons in this positive feedback loop can be targeted to regulate chronic pain and its concomitant symptoms, and whether gabaergic neurons of a class 3 neuronal VTA can also be targeted to regulate chronic pain and its concomitant symptoms, as demonstrated by virus injection, immunostaining, electrophysiology and behavioural (fig. 11). The results demonstrate that in SNI model mice, ACC glutamatergic neurons directly project and activate VTA gabaergic neurons, a class of inhibitory neurons that inhibit the dopaminergic neurons of the VTA zone to which it is directly connected. The inventors were able to effectively alleviate symptoms associated with nociception and anxiety-like symptoms in SNI model mice when inhibiting VTAGABA-competent neurons that received the projections of ACC glutamatergic neurons (fig. 11).
FIG. 11 a shows a schematic representation and a schematic representation of ACC and VTA region injection viruses (pAAV-CaMKII alpha-EGFP-2A-Cre-WPRE company: OBIO cargo number: CN870; pAOV-CAG-EGFP-2A-CRE company: OBIO cargo number: CN396; rAAV-EF1 alpha-DIO-EGFP-T2A-TK-WPRE-hGH pA company: brainVTA-0087; pAAV-GAD67-EGFP-2A-Cre-WPRE company: OBIO cargo number: H10062); b-D is an ACC region virus injection (pAAV-EF 1 alpha-DIO-hM 3D (Gqh) -EGFP-WPRE company: OBIO cat# H15959; pAAV-CaMKII alpha-EGFP-2A-Cre-WPRE company: OBIO cat# CN 870), electrophysiological recording the results of the increase in the frequency of VTA GABAergic neuron action potential emission and the decrease in the frequency of dopaminergic neuron action potential emission, respectively (c: n=16 cells/7 pairs of mice, two groups; D: n=13 cells/6 pairs of mice, two groups; p < 0.01); e-g shows that inhibiting viruses (rAAV-GAD 67-DIO-hM4D (Gi) -mCherry-WPRE-hGH pA company: brainVTA cat# PT-0608; rAAV-hSyn-CRE-EGFP-WPRE-hGH pA company: brainVTA cat# PT-1168) inhibit VTA GABAergic neuron activity receiving projection of ACC glutamatergic neurons, and can effectively relieve SNI mice from pain-sensitivity and accompanying anxiety-like behavior (n=10 pairs of mice; p < 0.05; p < 0.01). These results all indicate that VTA gabaergic neurons can also be an important regulatory target for chronic pain and its concomitant symptoms.
The results of the a-h correlation experiments in FIG. 11 (all expressed as the mean value of each group) are shown below:
/>
the above experimental results demonstrate that: VTA gabaergic neurons can be used as targets in the preparation of a medicament for the treatment of any of chronic pain and its concomitant symptoms. The medicament prepared from the VTA GABA energy neuron activity inhibitor can treat any one of chronic pain and accompanying symptoms.
Based on the application of the VTA DA energy neuron, the ACC glutamatergic neuron, the VTA GABA energy neuron or the dopamine D2 receptor serving as a drug target in relieving chronic pain and the accompanying symptoms thereof, the invention improves and treats the chronic pain diseases.
It is apparent that the above examples are given by way of illustration only and are not limiting of the embodiments. Other variations or modifications of the above teachings will be apparent to those of ordinary skill in the art. It is not necessary here nor is it exhaustive of all embodiments. While still being apparent from variations or modifications that may be made by those skilled in the art are within the scope of the invention.
Claims (22)
1. Use of a Ventral Tegmental Area (VTA) dopaminergic neuron, a anterior cingulate gyrus cortex (ACC) glutamatergic neuron, a VTA gamma-aminobutyric acid (GABA) ergic neuron, or a dopamine D2 receptor as a target in the manufacture of a medicament for the treatment of any one of chronic pain and its concomitant symptoms.
2. The use according to claim 1, wherein the chronic pain is selected from at least one of trigeminal neuralgia, parkinson's disease pain, sciatica, neuropathic pain, migraine and tumor pain.
3. The use according to claim 1, wherein the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear and cognitive impairment.
4. The use according to claim 1, wherein the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
Use of an enhancer of VTA DA-capable neuron activity, an inhibitor of ACC glutamatergic neuron activity or an inhibitor of VTA gabaergic neuron activity in the manufacture of a medicament for the treatment of any one of chronic pain and its concomitant symptoms.
6. The use according to claim 5, wherein the chronic pain is selected from at least one of trigeminal neuralgia, parkinsonian pain, sciatica, neuropathic pain, migraine and tumour pain.
7. The use according to claim 6, wherein the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear and cognitive impairment.
8. The use according to claim 5, wherein the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
Use of a D2 receptor agonist for inhibiting ACC glutamatergic neuronal activity projected by dopaminergic neurons in the ventral tegmental area in the manufacture of a product, preferably said D2 receptor agonist is selected from at least one of bromocriptine, a compound of formula (I) and salts thereof, cabergoline, pergolide, apo, pramipexole and Luo Paini Luo Zhong.
Wherein R1 is a C1-C6 alkyl group, preferably a C1-C3 alkyl group.
10. The use according to claim 9, wherein the D2 receptor agonist is an inorganic acid salt, preferably a hydrochloride salt, of a compound of formula (I), more preferably wherein the D2 receptor agonist is quinpirole or a salt thereof.
11. The use according to claim 9, wherein the product is a medicament for the treatment of any of chronic pain and its concomitant symptoms.
12. The use according to claim 10, wherein the chronic pain is selected from at least one of trigeminal neuralgia, parkinson's disease pain, sciatica, neuropathic pain, migraine and tumor pain.
13. The use according to claim 10, wherein the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear and cognitive impairment.
14. The use according to claim 10, wherein the medicament is a liquid formulation or a solid oral formulation; more preferably, the medicament is in the form of a suspension, elixir, solution, powder, granule, ointment, cream capsule or tablet; further preferably, the drug is an injection, a powder for injection or a tablet for injection.
Use of a D2 receptor antagonist for the preparation of a product acting on a VTA for enhancing the activity of dopaminergic neurons by releasing the self-inhibitory effect of the D2 receptor on dopaminergic neurons; preferably, the D2 receptor antagonist is selected from at least one of the compounds of formula (II) and salts thereof, sulpiride, metoclopramide, domperidone and etipride,
Wherein R1 is a C1-C6 alkyl group, preferably a C1-C3 alkyl group;
r2 and R3 are each independently optionally substituted C6-C12 aryl, preferably halogen substituted C6-C12 aryl, more preferably halogen substituted phenyl, further preferably halogen para-substituted phenyl, further preferably halogen is fluorine or chlorine.
16. The use according to claim 15, wherein the D2 receptor antagonist is haloperidol or a salt thereof.
17. The use according to claim 15, wherein the product is a medicament for the treatment of any of chronic pain and its concomitant symptoms.
18. The use according to claim 16, wherein the chronic pain is selected from at least one of trigeminal neuralgia, parkinson's disease pain, sciatica, neuropathic pain, migraine and tumor pain.
19. The use according to claim 16, wherein the concomitant symptoms are selected from at least one of anxiety, depression, aversion, fear and impairment of cognitive function.
20. A pharmaceutical composition comprising a D2 receptor agonist and a D2 receptor antagonist.
21. Use of the pharmaceutical composition of claim 20 for the manufacture of a medicament for the treatment of any one of chronic pain and its concomitant symptoms.
22. A kit comprising the pharmaceutical composition of claim 20, preferably the kit comprises instructions for administering the D2 receptor agonist and D2 receptor antagonist for use in the treatment of any of chronic pain and its concomitant symptoms.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311588201.3A CN117599181A (en) | 2023-11-27 | 2023-11-27 | Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311588201.3A CN117599181A (en) | 2023-11-27 | 2023-11-27 | Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117599181A true CN117599181A (en) | 2024-02-27 |
Family
ID=89943701
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311588201.3A Pending CN117599181A (en) | 2023-11-27 | 2023-11-27 | Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117599181A (en) |
-
2023
- 2023-11-27 CN CN202311588201.3A patent/CN117599181A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Mogenson et al. | From motivation to action: a review of dopaminergic regulation of limbic→ nucleus accumbens→ ventral pallidum→ pedunculopontine nucleus circuitries involved in limbic-motor integration | |
| Li et al. | Chronic stress exacerbates neuropathic pain via the integration of stress-affect–related information with nociceptive information in the central nucleus of the amygdala | |
| Olds | Hypothalamic substrates of reward | |
| Ryczko et al. | The multifunctional mesencephalic locomotor region | |
| Barchini et al. | Spinal segmental and supraspinal mechanisms underlying the pain-relieving effects of spinal cord stimulation: an experimental study in a rat model of neuropathy | |
| Vianna et al. | Dorsolateral and ventral regions of the periaqueductal gray matter are involved in distinct types of fear | |
| Shi et al. | High-frequency stimulation of the subthalamic nucleus reverses limb-use asymmetry in rats with unilateral 6-hydroxydopamine lesions | |
| Funk et al. | The rhythmic, transverse medullary slice preparation in respiratory neurobiology: contributions and caveats | |
| Negus et al. | Effects of kappa opioids in an assay of pain-depressed intracranial self-stimulation in rats | |
| Pei et al. | A partial trace amine‐associated receptor 1 agonist exhibits properties consistent with a methamphetamine substitution treatment | |
| Borelli et al. | Effects of acute and chronic fluoxetine and diazepam on freezing behavior induced by electrical stimulation of dorsolateral and lateral columns of the periaqueductal gray matter | |
| Gifuni et al. | Lesions of the lateral habenula dissociate the reward-enhancing and locomotor-stimulant effects of amphetamine | |
| Bolzoni et al. | Presynaptic and postsynaptic effects of local cathodal DC polarization within the spinal cord in anaesthetized animal preparations | |
| Stefani et al. | Subthalamic nucleus deep brain stimulation on motor-symptoms of Parkinson’s disease: focus on neurochemistry | |
| Hu et al. | Inhibition of P2X7R in the amygdala ameliorates symptoms of neuropathic pain after spared nerve injury in rats | |
| Estes et al. | Systemic oxytocin administration alters mesolimbic dopamine release in mice | |
| Jacob et al. | Chronic imipramine enhances 5-HT1A and 5-HT2 receptors-mediated inhibition of panic-like behavior in the rat dorsal periaqueductal gray | |
| Monti et al. | The effects of local microinjection of selective dopamine D1 and D2 receptor agonists and antagonists into the dorsal raphe nucleus on sleep and wakefulness in the rat | |
| Lazenka et al. | Sex differences in abuse-related neurochemical and behavioral effects of 3, 4-methylenedioxymethamphetamine (MDMA) in rats | |
| Lim et al. | Attenuation of fear-like response by escitalopram treatment after electrical stimulation of the midbrain dorsolateral periaqueductal gray | |
| De Luca-Vinhas et al. | Pharmacological assessment of the freezing, antinociception, and exploratory behavior organized in the ventrolateral periaqueductal gray | |
| Ferreira et al. | Infralimbic medial prefrontal cortex alters electroacupuncture effect in animals with neuropathic chronic pain | |
| Jourdain et al. | Potentiation of response to low doses of levodopa in MPTP-injected monkeys by chemical unilateral subthalamotomy | |
| CN117599181A (en) | Application of neuron as target in preparation of product for treating chronic pain and accompanying symptoms | |
| An et al. | Anxiolytic-like effect of mirtazapine mediates its effect in the median raphe nucleus |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination |