AT504705A1 - TREATMENT OF FEARING STATES - Google Patents

Description

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The present invention relates to formulations for the treatment and / or prevention of anxiety disorders as well as formulations for enhancing exploration behavior and perceptiveness.

In the field, various types of drugs are used in the treatment of anxiety disorders, including antidepressants and beta-blockers. These medications can be effective in controlling and relieving anxiety symptoms.

Many anxiety medications can also have unpleasant and unwanted side effects. These include tremors, headache, nausea, abdominal pain, dry mouth, weight gain, dizziness and sexual dysfunction. Anxiety medications can also be addictive, and long-term use can make withdrawal difficult.

For example, benzodiazepines, which are commonly prescribed to treat anxiety disorders, are extremely addictive to prolonged use. There may be drug tolerance within a few weeks. The consequence of this is that higher dosages are necessary to achieve the same relief as before. As addiction to anxiety medication becomes more prevalent, it becomes harder to stop taking it. If the administration of benzodiazepines is stopped abruptly, it can lead to severe withdrawal symptoms such as restlessness, insomnia and renewed anxiety. Deprivation also works when selective serotonin reuptake inhibitors are administered during the treatment of anxiety disorders. Excessive withdrawal may cause symptoms such as dizziness, nausea, tiredness and memory problems. Great anxiety and depression are also common in withdrawal symptoms.

Cognitive enhancers (CE) are compounds that increase the final level of performance (1). The development of such substances is of clinical importance for increasing cognitive performance or treating cognitive dysfunction associated with aging, neurodegenerative diseases and psychiatric disorders. A number of compounds have been reported as potential memory or cognitive CEs (see, e.g., Table A), and some have already been evaluated in clinical trials (2-4). The modulation of several receptor systems was investigated.

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Table A: Potential Cognitive Enhancers DRUG LABYRINTH / ANIMAL LIT. D-Cycloserine Partial agonist NMDA R-associated glycine site T-labyrinth abandonment: (Male Long-Evans) rats 15 Passive Avoidance Task: (Long-Evans) Rats Thirst-motivated Lineage Maze Lera Task (Male Swiss) Mice 13 Puebstoßvermeidungsaufgäbe. Mice 19 Arm Radial Labyrinth (DNMS). (Male Black-Wistar) Rats 17,47, 48 Wet Labyrinth Challenge (Reference Memory) Rats 14, 20 RS67333 Agonist 5-HT4 Receptor Morris-Nasser Labyrinth: (Sprague-Dawley) Rats; rats treated with atropine 17, 41, 42 SIB-1553A Agonist β-4 nicotine receptor Delayed Non-Matching to Place: C57BL / 6 mice 26, 27 T-Labyrinth: C57BL / 6 mice Morris Water Maze: Old Fisher rats SGS742 GABAb Antagonist Passive Avoidance Test: Male Mice 36, 4 Social Learning Task: Male Spraque-Dawley Rats 36 8-Arm Radial Labyrinth Retrieving: Long-Evans Rats 32 N-methyl-D-aspartate (NMDA) receptors are exemplified in U.S. Pat Nerve pathways are involved in learning and memory formation, and several pharmacological studies that have shown that pre-workout antagonists of the NMDA receptor complex disrupt certain types of learning and memory (5-7) support this conclusion.

An object of the present invention is to provide new medicaments and therapies for the treatment of anxiety disorders. Another object of the present invention is to provide formulations that enhance exploration behavior and perceptiveness.

Therefore, the present invention relates to the use of

I FOLLOW-UP 3 3 • • • • • D-Cycloserine (DCS) for the manufacture of a medicament for the treatment or prevention of anxiety disorders. D-cycloserine (DCS) was originally used as an antibiotic to treat tuberculosis and was later characterized as a partial agonist at the NMDA receptor-glycine binding site (8-11). DCS appears to be potent CE in lower doses than required for antibacterial activity (12-14). It has been reported as a performance enhancer in various learning and memory assessment tasks (15-17). In doses used in most studies, DCS has no appreciable effect on the motivational state or motor skills of the experimental animals, suggesting that the effects of DCS are most likely due to specific effects on learning and memory (18).

Most researchers have observed that administration of DCS to rats and mice promotes spatial learning (12, 14). Retention of a thirst-motivated linear labyrinth task was improved by administering 10 mg / kg, 20 mg / kg, or 80 mg / kg DCS to mice immediately after exercise. In addition, a dose of 3 mg / kg DCS administered 20 minutes before training facilitated learning this task. Post-training acute post-exercise injections did not facilitate retention when the mice were pretreated with DCS (3 mg / kg) in the maze for 15 days prior to training. Not only did the study show that the administration of DCS improved consolidation and memory recall, but also that desensitization could occur when the drug was given chronic action (16). Trained young mice injected with 20 mg / kg DCS also showed increased memory retention, as demonstrated by an electroshock-motivated T-maze task (19). Surprisingly, it has been shown that the administration of D-cycloserine (DCS) allows the treatment and / or prevention of anxiety disorders. "Anxiety disorders " As used herein, refer to disorders that show several different forms of abnormal, pathological anxiety, fear, phobias, and nervous conditions that may occur suddenly or gradually over a period of several years and may prevent or prevent a continuation of the normal daily routine. 1 n

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Fear and fear are ubiquitous sensations. Fear is defined as an emotional and physiological response to a perceived external threat. Fear is an unpleasant emotional state whose origins are not easily identified. It is often accompanied by physiological symptoms that can lead to fatigue or fatigue. Because the fear of perceived threats causes similar unpleasant mental and physical changes, patients use the expressions fear and anxiety alternately. It does not necessarily distinguish between fear and fear. What is important, however, is the distinction between the various anxiety disorders, as an accurate diagnosis tends to result in more effective treatment and better prognosis.

According to a preferred embodiment of the present invention, the anxiety disorder is selected from the group consisting of generalized anxiety disorder, panic disorder, agoraphobia, phobias, social anxiety disorder, obsessive-compulsive disorder and posttraumatic stress disorder.

General anxiety disorder is an anxiety disorder characterized by excessive and uncontrollable worries about daily things. Frequency, intensity and duration of discomfort are disproportionate to the actual source of concern, and such worries often hinder daily routine. About 5% of the total population is affected. Symptoms of a common anxiety disorder are tension, dread, restlessness, hyperactivity, pondering, fear, and contemplation. These symptoms must be consistent and occur at least every other day and last for at least 6 months.

About 30% of the causes of a generalized anxiety disorder are inherited, but certain characteristics make people more vulnerable to it. People with general nervousness, depression, inability to be frustrated and feeling narrowed are more likely to be found among patients with generalized anxiety disorder. In adolescence, generalized anxiety disorders often result in lower social support, academic underachievement, underemployment, substance use, and a high probability of acquiring other psychiatric disorders.

General anxiety disorder becomes conventional with selective serotonin and serotonin norepinephrine reuptake inhibitors

I REACHED 5 (SSRI or SNRI). Preferred clinically used inhibitors are fluvoxamine, sertraline, paroxetine, citalopram, escitalopram, venlafaxine, pregabalin, buspirone and benzodiazepines such as diazepam and alprazolam.

People who suffer from panic disorder usually experience a series of intense episodes of extreme anxiety, also known as panic attacks. A panic event can be triggered by a particularly stressful situation or occur for no particular reason. Such events usually take several minutes. 35% of all panic disorder sufferers also have agoraphobia.

For the treatment of panic disorders selective serotonin reuptake inhibitors, monoamine oxidase inhibitors and benzodiazepines may be administered. A medicament for use in the treatment of panic disorder may include one or more of the following: alprazolam, bromazepam, chlordiazepoxide, clobazam, clonazepam, clorazepate, diazepam, halazepam, paxipam, ketazolam, lorazepam, medazepam, nordazepam, oxazepam, prazepam, harmalin, Iproclozide, iproniazid, isocarboxazid, nialamide, phen-enzine, selegiline, toloxaton, tranylcypromine, citalopram, escita-lopramoxalate, fluoxetine, fluvoxamine maleate, paroxetine, sertraline, dapoxetine, isocarboxazid, moclobemide, phenelzine, tranylcypromine, selegiline, nialamide, iproniazid, iprocloth and toloxatone.

Agoraphobia is an anxiety disorder that consists primarily of the fear of a difficult or unpleasant situation, from which there is no escape for the sufferer.

People with agoraphobia can experience severe panic attacks in situations where they feel trapped, insecure, out of control, or too far from their personal comfort zone. In severe cases, it can occur that Agoraphobians not only retreat to their homes but into one or two rooms, or become bedridden or hermit. They often react extremely sensitively to their own body sensations and overreact in the subconscious mind to completely normal occurrences.

People who suffer from agoraphobia can be treated with anti-anxiety and / or antidepressant medications.

A phobia is a strong lingering fear of situations, objects, activities or people. The main symptom of this disorder consists in excessive, irrational

FOLLOWING 6 demanding longing to avoid the dreaded subject. People with phobias may be treated with anti-anxiety and / or antidepressant medications.

Social anxiety is an experience of fear, concern, or unrest about social situations and the ability to be judged by others. Physical symptoms, which often go hand in hand with social anxiety disorders, include excessive redness, sweating (hyperhydrosis), tremors, nausea and stammering. In case of great fear and anxiety, it can also lead to panic attacks. People suffering from social anxiety disorders may be treated with antidepressants such as monoamine oxidase inhibitors, benzodiazepines and / or selective serotonin reuptake inhibitors.

Obsessive-compulsive disorders are a type of anxiety disorder characterized primarily by obsession and / or obsession. Obsession manifests itself in embarrassing, repetitive, annoying thoughts or images that the individual often sees as meaningless. Obsessions are repetitive behaviors that a person feels pressured or forced to rid themselves of. People suffering from obsessive-compulsive disorder may be treated with medications containing selective serotonin reuptake inhibitors, especially paroxetine, sertraline, fluoxetine and fluvoxamine, tricyclic antidepressants, especially clomipramine, gabapen-tin, lamotrigine, olanzapine and risperidone, include.

Posttraumatic stress disorder is the term for a particular psychological sequelae after the impact of a stressful experience or the confrontation with a stressful experience that the person experiences as highly traumatic. Such experiences include actual or threatened death, serious bodily injury or threat to physical and / or mental integrity. Symptoms of this disorder may include nightmares, flashbacks, emotional detachment or dulling of feelings, insomnia, avoidance of memory aids, and extreme distress in the face of memory, irritability, hypervigilance, memory loss, and excessive anxiety reaction, clinical depression and anxiety, loss of appetite. Posttraumatic stress disorder can be treated with antidepressants.

According to another preferred embodiment of the present

According to the invention, the medicament is formulated for oral or parenteral administration, in particular for intravenous, intramuscular, subcutaneous, transdermal, transmucosal or inhalatory administration, wherein the dosage forms are preferably selected from the group consisting of food supplement, aerosol , Capsule, powder, solution, suspension, tablet, injection, infusion, gel, paste and transdermal patch.

Depending on the route of administration and dosage form, the medicament preferably comprises at least one pharmaceutically acceptable excipient, carrier and / or diluent.

Methods and apparatus for producing medicaments in a particular dosage form are known to those skilled in the art (see, e.g., "Handbook of Pharmaceutical Manufacturing Formulations").

Niazi KS, ISBN: 0849317525, CRC Press Inc .; " Handbook of Pharmaceutical Excipients " Rowe RC, Sheskey PJ, Owen SC, ISBN: 1582120587, APhA Publications).

Especially preferred is the formulation of medicaments of the present invention for oral administration, especially as tablets or capsules.

The medicament of the present invention preferably comprises, in addition to DCS, at least one further active ingredient, preferably at least one further anxiolytic agent selected from the group consisting of D-cycloserine, alprazolam, bromazepam, chlordiazep-oxide, clobazam, clonazepam, clorazepate, diazepam, halazepam, paxipam, ketazolam , Lorazepam, medazepam, nordazepam, oxazepam, prazepam, harmalin, iprocloth, iproniazid, isocarboxazid, nialamide, phenelzine, selegiline, toloxaton, tranylcypromine, citalopram, escitalopram oxalate, fluoxetine, fluvoxamine maleate, paroxetine, sertraline, dapoxetine, isocarboxazid, moclobemide, phenelzine , Tranylcypromine, selegiline, nialamide, ipronocid, iprocloth, toloxaton, a barbiturate and meprobamate.

To further increase the efficacy of the medicament of the invention, this medicament may further comprise at least one other anxiolytic. Particularly suitable anxiolytics are mentioned above. The at least one further anxiolytic may also be selected depending on the anxiety disorder to be treated (see above).

According to a preferred embodiment of the present invention, D-cycloserine is administered in the amounts already established for this compound, in particular in an amount

From 0.5 to 50 mg / kg of body weight, preferably 1 to 40 mg / kg of body weight, more preferably 1.5 to 30 mg / kg body weight, especially 2 to 20 mg / kg body weight. DCS already shows beneficial properties in the treatment of anxiety disorders at doses of 0.5 mg / kg body weight.

Another aspect of the present invention relates to the use of D-cycloserine (DCS) in a formulation for enhancing exploration, comprehension and receptivity. Surprisingly, it has been found that the administration of DCS to an individual and to animals enhances the exploration behavior, comprehension and receptivity of that individual or animals. Therefore, DCS may be administered to an individual or animal to increase exploration, comprehension, and capacity. Because of these features, DCS can be administered not only to people suffering from a disease, but also, and most preferably, to individuals and / or animals to increase their mental and intellectual capacity.

The formulation according to the present invention can therefore be used in various fields of application.

According to a preferred embodiment of the present invention, D-cycloserine is administered for oral or parenteral administration, in particular for intravenous, intramuscular, subcutaneous, transdermal, transmucosal or inhalatory administration, preferably as a nutritional supplement, aerosol, capsule, powder, solution, suspension, tablet, injection, Infusion, gel, paste or transdermal patch, formulated.

Such formulations and dosage forms preferably comprise at least one pharmaceutically acceptable excipient, carrier and / or diluent.

Methods and apparatus for producing medicaments in a particular dosage form are known to those skilled in the art (see, e.g., "Handbook of Pharmaceutical Manufacturing Formulations").

Niazi KS, ISBN: 0849317525, CRC Press Inc .; " Handbook of Pharmaceutical Excipients " Rowe RC, Sheskey PJ, Owen SC, ISBN: 1582120587, APhA Publications).

FIELD D-cycloserine is preferably administered in an amount of 0.5 to 50 mg / kg, preferably 1 to 40 mg / kg, more preferably 1.5 to 30 mg / kg, especially 2 to 20 mg / kg. - 9 • · ··· * • ·· • '··

The present invention is further illustrated in the following figures and examples, without being limited thereto.

Figure 1 shows the escape latency (a) and spatial preference patterns during a probe test (b) on DCS-treated C57BL / 6J mice compared to vehicle-treated C57BL / 6J mice. The asterisks indicate the significant differences in drug-treated mice compared to the control or reference drugs (SGS742, SIB1553A, or RS6733) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, *** p £ 0.001).

Figure 2 shows the escape latency (a) and spatial preference patterns during a probe test (b) on DCS treated DBA / 2 mice compared to vehicle treated DBA / 2 mice. The asterisks indicate the significant differences between drug-treated mice compared to the control and reference drugs (SGS742, SIB1553A and RS6733) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, P <0.001).

Figure 3 shows the escape latency (a), range (b) and spatial preference patterns during a probe test (c) on DCS-treated C57BL / 6J mice compared to vehicle-treated C57BL / 6J mice. The asterisks indicate the significant differences between drug-treated mice compared to the control and reference drugs (SGS742, SIB1553A and RS6733) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, *** P £ 0.001).

Figure 4 shows the average speed during the learning and consolidation phase in MWM for DCS treated DBA / 2 (a) and C57BL / 6J mice compared to their control groups. The asterisks indicate the significant differences between drug-treated mice compared to the control and reference drugs (SGS742, SIB1553A and RS6733) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, *** P £ 0.001).

EXAMPLE 1. Materials

Male C57BL / 6J and DBA / 2 mice were used at the age of 10-14 weeks. The mice were reared in the Core Unit of Biomedical Research, Department of Laboratory Animal Science and Genetics, Medical University of Vienna and in

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Held macroion cages that were filled with autoclaved wood chips. An autoclaved standard rodent diet (Altromin 1314ff) and acidified or pH 3 tap water or bottled water was available ad libitum. The room temperature was 22 + 1 ° C and the relative humidity 50 + 10%. The aeration with 100% fresh air gave an air exchange rate of 15 times per hour. The room was lit with artificial light of about 200 lx in 2 m from 5 to 17 o'clock. Between 8:00 and 13:00, behavioral tests were conducted.

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Animals were treated intraperitoneally (i.p.) before daily testing. The mice were divided into four treatment and one control group (vehicle group) (0.9% NaCl solution). Doses and injection times were chosen based on research in the literature (see Table 1).

Table 1: Drugs used in the experiments and the corresponding injection times and doses on the basis of literature research. ARZNEIEN INJECTION TIME CAN LIT. DCS 30 min before testing 10 mg / kg [14] DCS was purchased from Sigma-Aldrich (Austria). 2. Procedure for behavioral training

Behavioral Observation Screen: a) Basic Neurological and Physiological Observation Assessment: The procedure follows the arrangement of Irwin (44). A test battery was used to diagnose defects in gait or posture, changes in muscle tone, gripping power, visual acuity, and temperature. To complete the assessment, vital reflexes were evaluated. In addition, abnormal behavior, fear, irritability, aggression, excitability, salivation, lacrimation, urination, and defecation were recorded during the manipulations. b) Rotarod: In the Rotarod test (Rota Rod "Economex", Columbus Instruments, USA), balance and coordination are tested by means of a rotating roller which is accelerated from 4 to 40 rpm over 5 minutes. The time was recorded, too

FOLLOWING - 11 ····················································································································· Each animal received three pre-training tests. Afterwards, each mouse completed three more consecutive tests, and the analysis was then used the longest time on the roller (45). c) Elevated-Plus-Maze (EPM): A paradigm used to study anxiety in mice. The animals were observed for 5 minutes on a labyrinth consisting of 4 arms (each 30 cm long and 5 cm wide) fixed at a height of 54 cm. The arms are connected by a 5 cm x 5 cm wide central area. Two arms contain 15 cm high side and end walls (= closed arm). It evaluated standard parameters reflecting anxiety-related behavior (ie, entries into the closed arms, length of stay in the closed arms, frequency of defecation) (46). d) Open Field (OF): Using a video surveillance system consisting of a video camcorder coupled to a TiBeSplit computer location system, mice were observed in an arena (40 cm x 40 cm long, with 70 cm high walls) for 10 minutes , The individual mice were placed in the middle of the chamber for each experiment. The default parameters for motion activity (ie the total distance covered, the average speed, the extent of large movements, the extent of local movements, rest time, the frequency of spontaneous directional changes) and exploration behaviors (ie rearing, center crosses, length of stay in the middle ) (46).

Morr is VJater Maze learning task

The Morris Water Labyrinth MWM consisted of a round basin (122 cm diameter, walls 76 cm deep) in which the mice were trained to escape the water by swimming to a hidden platform (1.5 m below the surface of the water) whose location could be located only by using remote, attached to the room walls outside the labyrinth orientation aids. The water temperature was kept at 21 + 1 ° C.

The pelvis was divided into four quadrants using a computerized location / image analyzer system (video camcorder: Sony CVX-V18NSP coupled to a computer location system: TiBeSplit). The platform was located in the middle of the SW quadrant

POSITIONED and remained in one and the same place throughout the experiment.

The spatial learning task consisted of 4 training attempts per day and 4 training days. The mice were released from the four compass directions (NO, NW, SW, and SO, in that order) with their heads toward the pelvic wall, and were allowed to swim for 120 seconds looking for the platform. If the mice did not locate the platform after 120 s, the animals were placed manually on the platform and left there for 30 s. Each animal was then returned to its cage for 10 minutes before starting the next trial. On the first day of training, the mice were given a training session for acclimatization in the water labyrinth of 30 s.

One day after the last day of training, the subjects were subjected to a probe test with the platform removed. The mouse was released from the NO start point and allowed to float freely for 60 seconds. The path taken by the mouse swimming was tracked and analyzed for the ratio swim time and / or distance in each quadrant of the pool, and the swimming speed was recorded. 3. Experimental setup

First, the Open Field Test, the Elevated Plus Maze Experiment, the Neurological Observation Battery, and the Rotarod Assessment were conducted to exclude behavioral influences and confounding factors due to CEs (such as anxiety related behaviors).

The order of the tests was designed so that the procedures most likely to be affected by pretreatments were performed first (45): EPM, OF, battery of neurological observations and RR. The drugs were given before the tests. One week later, after completion of the behavioral observation screen, all mice were scored in MWM to demonstrate the effect of CEs on memory formation. 4. Statistical analysis:

A statistical analysis to show differences between the groups was made using the unpaired Student's t-test. Where data precluded a principal assumption of parametric distribution, a non-parametric Mann-Whitney U test was performed. Where appropriate, Fisher's exact test and chi-square test were also used.

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In all cases, a probability level P &lt; 0.05 considered statistically significant. All calculations were done using GraphPad InStat version 3.00 for Windows (GraphPad Software, USA). 5. Results a) Elevated Plus Maze

As shown in Table 2a, the total distance traveled in closed arms in C57BL / 6J inbred mice was significantly reduced by DCS compared to the vehicle control group. The length of stay in closed arms was significantly lower in both treated groups, DCS and SGS742, compared to the control group. Rest time was lower only in the DCS-treated mice.

Table 2a: Results of the EPM test. The asterisks show the significant differences between drug-treated mice (C57BL / 6J and DBA / 2) compared to the control group (0.9%).

NaCl solution) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, P ^ 0.001). C57BL NaCl (n = 12) DCS (n-16) Closed arm (CA) left 11.3 * 3.2 11.6 * 3.8 To the covered distance in CA (m) 6.1 ± 1.8 4.9 ± 1.7 * Duration of stay in CA (s) 199.5 * 51.8 153.0 ± 49.3 * Total covered distance (m) 8.1 ± 1.2 8.7 ± 1.6 Resting time (s) 66.6 * 5 , 3 62.1 ± 6.4 * extent of major movements (%) 4.5 ± 1.9 4.3 ± 2.7 defecation 0.4 ± 0.1 0.4 ± 1.1

NaCI DCS DBA (n = 11) (n = 16) Closed arm (CA) left 8.0 ± 5.0 7.6 ± 3.1 Distance traveled in CA (m) 6.5 ± 2.1 6.5 ± 1.4 Duration of stay CA (s) 218.2 ± 54.8 232.6 * 31.5 Total covered distance (m) 7.9 ± 1.8 7.4 ± 1.0 Rest period 71.0 ± 6.8 73.7 * 3.5 extent of large movements (%) 5.7 ± 3.1 5.3 ± 1.8 Stool emptying 0.8 ± 1.0 1 ± 1.2 FOLLOWING lb) Open-field

As shown in Table 2b, the total distance traveled was significantly reduced in DCS-DBA mice. DCS significantly reduced resting time and length of stay in the limb and significantly increased the number of midcrossings, the frequency of spontaneous changes in direction, and the extent of large movements. For C57BL / 6J, DCS significantly increased the number of center intersections and the extent of rearing and cleaning.

Table 2b: Results of the OF test. The asterisks show the significant differences between drug-treated mice (C57BL / 6J and DBA / 2) compared to the control group (0.9% NaCl solution) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, P ^ O, 001).

SUBSEQUENT

NaCl DCS C57BL (n-12) (n * 16) Total distance traveled (m) 33.3 ± 6.0 34.8 ± 6.5 Rest period 46.6 ± 5.4 46.4 ± 6.8 Average speed (m / s) 0.05 ± 0.01 0.06 ± 0.01 Number of center intersections 7.5 ± 4.5 11.6 ± 4.5 * Time of snout / rearing 90.8 ± 39.5 128.5 ± 38.8 ** Frequency of spontaneous changes of direction 43.1 ± 11.8 47.7 ± 10.0 Duration of stay in limitation 56.3 ± 11.75 58.1 ± 10.7 Extent of large movements (%): Movement speed> 0.10 m / s (% of the total observation time) 23.0 ± 4.6 24.1 ± 4.9 Extent of local movement (%): Movement speed 0.03m / s <x <0.10m / s (% of total observation time) 30.4 ± 2.2 29.5 ± 2.5 defecation 1.3 ± 1.7 0.7 ± 0.8

NaCl DCS DBA <n = 11) (n = 16) Total distance traveled (m) 27.9 ± 7.7 34.7 ± 5.6 * Resting time 60.0 ± 11.9 51.7 ± 3.6 * * Average speed (m / s) 0.05 ± 0.01 0.06 ± 0.01 * number of center intersections 2.7 ± 3.1 7.8 ± 3.5 *** extent of clearing and cleaning 99.2 ± 25.1 99.6 ± 25.5 Frequency of spontaneous changes in direction 29.3 ± 11.4 44.8 ± 11.8 ** Duration of stay in the limit 77.2 ± 11.3 67.4 ± 8.1 * * Extent of large movements (%): movement speed> 0.10 m / s (% of the total observation time) 17.1 ± 6.0 23.2 ± 4.0 ** amount of local movement (%): movement speed 0, 03m / s <x <0.10m / s (% of total observation time) 23.0 ± 6.2 25.1 ± 1.9 defecation 1.7 ± 1.7 2.3 ± 1.4 c) observation battery

The amount of results is given in Tables 2c.I and 2c.II and will not be discussed here.

Table 2c.I: Results of the primary behavioral observation screen in C57BL / 6J mice. Scores (mean +/- standard deviation) as described by Irwin et al. Evaluated in 1968: a higher score means more (better, higher) activity (performance, response), with the exception of gait, wire maneuver and righting reflex, which perform better at lower scores. Some parameters are rated as follows: +/- (presence / absence)

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C57BL NaCl (n = 12) DCS (n-16) A. Behavior 1. Spontaneous activity Body position 5.5 ± 2.3 5.9 ± 2.4 Activity 1, 8 ± 2.6 2.5 ± 2 Bizarre behavior 0 0 2. Motor affective responses Vestibular decline 2.8 ± 1.7 2.9 ± 1.3 Transfer excitation 2.9 ± 1.2 3.4 ± 0.5 Spatial locomotion 4,2 ± 1,4 4,8 ± 1,1 Attention 3,7 ± 1,4 5,8 ± 1,8 ** Provoked biting 3 ± 2 3,6 ± 2,0 Finger approach 2,8 ± 2, 5 2.5 ± 2.1 Finger retraction 1.5 ± 2.3 1.6 ± 2.2 Cliff avoidance 12/0 15/1 Vocalization 1.25 ± 1.1 1.8 ± 0.8 Urination 10/2 8 / 8 3. Sensorimotor Reaction Fright reaction 3.2 ± 2.8 5.3 ± 2.8 * Pinnareflex 1.5 ± 2.8 1 ± 2.2 Cornea 2.3 ± 1.7 2.8 ± 1.2 Toes pinch 1 ± 1.8 0.3 ± 0.7 Visual placement 4.5 ± 1.2 4.1 ± 0.9 Tailpain 0.1 ± 0.3 0.5 ± 1.0 B. Neurology 1. Stance Limb rotation 1.7 ± 0.8 1.1 ± 0.8 Pelvic lift 3.3 ± 1.1 4.3 ± 1 * Tail lift 2.7 ± 0.8 2.5 ± 0, 9 2. Mu Skeletal muscle tone 5,8 ± 1,3 5,8 ± 1,3 Gripping force 4,8 ± 1,8 3,6 ± 1,3 * Limb tone 3,3 ± 1,5 4,4 ± 1,6 * Abdominal tone 5 , 2 ± 1.8 5.1 ± 1.7 Wire maneuver 2.7 ± 2.3 3.6 ± 2.2 3. Balance and Gear Gait 0/12 1/15 Raise Reflex 0 0

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NaCI DCS C57BL <n = 12) (n®16) 4. CNS excitation ticks / twitch 0 0 C. AUTONOM 1. Eyes Eyelid closure 0 0.4 ± 1.5 Proprioception 5.9 ± 0.7 6 ± 0 2. Secretion and excretion Stool emptying 2.8 ± 1.3 1.9 ± 1.4 Diarrhea 1/11 0/16 Urination 10/2 8/8

Table 2c.II: Results of the primary behavioral observation screen in DBA mice. Scores (mean +/- standard deviation) as described by Irwin et al. Evaluated in 1968: a higher score means more (better, higher) activity (performance, response), with the exception of gait, wire maneuver and righting reflex, which perform better at lower scores. Some parameters are scored as follows: +/- (presence / absence) DBA NaCl (n = 11) DCS (n = 16) A. Behavior 1. Spontaneous activity Body position 5.7 ± 2.2 6.4 ± 1.8 Activity 2 ± 1.8 1.5 ± 1.6 Bizarre Behavior 2. Motivated Affective Reactions Vestibular Waste 0.7 ± 1.3 1.8 ± 1.3 * Transfer Excitement 2.9 ± 1.3 2.6 ± 1.5 Spatial Movement 3, 9 ± 1,9 3,7 ± 1,2 Berethrungsflucht 3,8 ± 1,1 4,3 ± 1,9 Provoked biting 1,5 ± 1,3 2,8 ± 2,3 Finger approach 2,6 ± 2, 5 3.6 ± 2.9 Fingertips 2.2 ± 2.6 1.3 ± 1.9 Cliff Avoidance 10/1 16/0 Vocalization 1.7 ± 0.9 2.4 ± 1.9 Urine

FOLLOW-UP 18 ΦΦ Φ Φ Φ Φ Φ Φ Φ ΦΦ ΦΦ Φ Φ Φ Φ ΦΦΦΦ ΦΦ Φ Φ · Φ Φ · Φ ΦΦ ΦΦ ΦΦΦΦ ΦΦ ΦΦ ·· ΦΦ DBA NaCl (n * 11) DCS <η = 1β) 3. Sensomotoric reaction Bock reaction 2.6 ± 1.8 2.1 ± 2.3 Pinnareflex 0 0.9 ± 2.1 Cornea 2.6 ± 1.1 2.6 ± 1.4 Toes pinch 1 , 5 ± 2.0 1.3 ± 1.4 Visual placement 4.4 ± 1.2 4.5 ± 0.9 Tail pincer 0.9 ± 1.6 1.9 ± 2.8 B. Neurology 1. Posture Rotation of the Limbs 1.3 ± 0.5 1.6 ± 0.5 Lifting the pelvis 3.5 ± 1.3 3.6 ± 1.1 Lifting the tail 1.8 ± 0.6 2 ± 0.4 2. Muscle tone Body tone 6.1 ± 0.3 6.8 ± 0.9 * Grasping force 4.4 ± 1.2 5.6 ± 1.6 * Limb tone 2.7 ± 2.0 3.7 ± 1.7 Abdominal tone 5, 5 ± 1.3 5.8 ± 1.4 Wire maneuvers 0.7 ± 1.9 1.8 ± 2.9 3. Balance and gait Gait 0/11 1/15 Erectile reflex 0 0 4. CNS excitation Taut / twitching 0 0 C. AUTONOM 1. Eyes Eyelid closure 0 0 Proprioception 6 ± 0.8 5.9 ± 0.9 2. Secretion and excretion Diarr ur 2/9 7/9 Urinating 2/9 3/13 defecation 12,6 ± 1,7 2,5 ± 1,8 d) Rotarod

Rotarod performance did not change significantly in DBA-treated mice compared to the control group.

Table 2d: results of the Rotarod, the longest time on the

SUBSEQUENT

Roller was used for the analysis. The asterisks show the significant differences between drug-treated mice (C57BL / 6J and DBA / 2) compared to the control group (0.9% NaCl solution) (t-Student test and / or Mann-Whitney U test; * P <0.05, ** P <0.01, P <0.001).

Maximum Test NaCl DCS C57BL 13.3 ± 7.8 14.75 ± 7.8 DBA 8.7 ± 7.3 12.6 ± 6.1 e) Morris Water Maze Challenge

Figures 1 and 2 show the spatial learning curves representing the mean + SD latencies in searching the underwater platform per mouse strain during the 4 days of learning training and the length of stay in the target quadrant during the probe trial.

In the C57BL / 6J inbred strain, no significant difference was found between the control and treatment groups during the first 2 days of training. On day 3, D-cycloserine terminated the experiment significantly faster (within 45.1 ± 35.8 and 40.1 ± 27.1, respectively) than the control group (52.6 ± 18.0). On day 4, however, no significant performance differences were observed (see FIG. 1a). During the consolidation phase (Figure lb), the length of stay of C57BL / 6J in the target quadrant was measured.

The DBA / 2 mice (Figure 2a) learned between the groups without significance during the first two days. On day 3, treated mice (D-cycloserine: 53.3 ± 20.7) took significantly longer to reach the platform than the control group (33.9 + 26.3). Again, no significant latency difference was seen in achieving the goal on day 4.

In the probe experiment, mice treated with D-cycloserine (12.3 ± 5.5) performed worse than the control group (16.2 + 1.5) because they spent significantly less time in the target quadrant (see Figure 2b).

When comparing the latency to reach the platform on the training days between DBA / 2 and C57BL / 6J, the control groups showed no difference on day 1, 2 and 4. However, on day 3, the C57BL / 6J mice (52.6 ± 18 , 0) significantly longer,

NACHGER9CHT - 20 • · · · · · · · · · · · · · · · · ································································································································································································

In contrast, the C57BL / 6J mice (19,113,8) gave significantly better probe performance than the DBA / 2 mice (16,211.5) (see Figure 3).

The average speed also changed significantly over the course of the training days, and the probe trial was comparable to the control groups in both strains (see Figure 4). 6. Summary

The main finding of the study is that CEs exert remarkable stem-related effects on behavioral, cognitive and neurological functions. Furthermore, evidence was provided for the presence of probable neurotoxic effects.

Decades ago, DCS was described as a potential cognitive enhancer that improved rodent spatial memory. In the case of Sprague-Dawley rats, daily intraperitoneal administration of DCS increased learning in MWM (17), decreased latency in a water labyrinth at 4 MO aging rates (14), and a smaller number of errors in adult Wistar rats observed an arm radial labyrinth task (55). Aura and co-workers showed that intraperitoneal administration of DCS improved learning in old but not young Han: Wistar rats in an MWM task (56). Multiple doses failed to improve learning and memory in male Han: Wistar rats using the MWM paradigm (57). Thus, no consistent CE effect of DCS was observed, and this may well be due to the different rat strains, age, test systems.

In mice, post-exercise administration of DCS increased memory retention in a thirst-motivated linear labyrinth in Swiss Webster mice (16), but no other strains or test systems have been used. In the present study, no effect on spatial memory in MWM was observed for C57BL / 6J. In DBA / 2, however, consolidation, expressed as reduced length of stay in the MWM's destination quadrant, was impaired, and the learning curve showed disturbed learning at least on the third day of training.

Behavioral and neurological side effects should not be taken as the cause of poor performance in MWM in DBA / 2 mice, and again, the underlying

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• Μ ·

Neuropathology or different strains have been responsible for the failure of DCS treatment. Observations of an extended total distance traveled, the number of center intersections, the frequency of spontaneous directional changes, and the magnitude of large movements may indicate hyperactivity that may have contributed to cognitive decline. Findings from this study do not refute the reported CE effects on mouse spatial memory per se, but a failure to improve spatial memory in C57BL / 6J clearly demonstrates that more work is required before DCS is credited with a clear CE effect can be. Behavioral effects in terms of reduced anxiety-related behavior as shown in the OF and EPM (Table 2a, b) confirm earlier findings of anxiolytic effects in low-anxiety rats (58).

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SUBSEQUENT

Claims (12)

1. A use of D-cycloserine (DCS) for the manufacture of a medicament for the treatment or prevention of anxiety disorders. 2. Use according to claim 1, characterized in that the anxiety disorder is selected from the group consisting of panic disorder, agoraphobia, phobias, social anxiety disorder, obsessivkompulsiver disorder and post-traumatic stress disorder. 3. Use according to claim 1 or 2, characterized in that the medicament is formulated for oral or parenteral administration, in particular intravenous, intramuscular, subcutaneous, transdermal, transmucosal or inhalatory administration. 4. Use according to any one of claims 1 to 3, characterized in that the medicament is formulated as a nutritional supplement, aerosol, capsule, powder, solution, suspension, tablet, injection, infusion, gel, paste or transdermal patch. Use according to any one of claims 1 to 4, characterized in that the medicament comprises at least one pharmaceutically acceptable excipient, carrier and / or diluent. 6. Use according to one of claims 1 to 5, characterized in that the medicament at least one further active substance ff, preferably at least one further anxiolytic agent selected from the group consisting of alprazolam, bromazepam, chlordiazepoxide, clobazam, clonazepam, clorazepate, diazepam, halazepam, Paxipam, Ketazolam, Lorazepam, Medazepam, Nordazepam, Oxazepam, Prazepam, Harmalin, Iprocloid, Iproniazid, Isocarboxazid, Nialamide, Phenelzine, Selegiline, Toloxaton, Tranylcypromine, Citalopram, Escitalopramoxalate, Fluoxetine, Fluoxamine Maleate, Paroxetine, Sertraline, Dapoxetine, Isocarboxazide, Moclobemide, Phenelzine, tranylcypromine, selegiline, nialamide, ipronocid, iprocloth, toloxaton, a barbiturate and meprobamate. 7. Use according to any one of claims 1 to 6, characterized in that D-cycloserine in an amount of 0.5 to 50 mg / kg, REPLACED [preferably 1 to 40 mg / kg, more preferably 1.5 to 30 mg / kg , in particular 2 to 20 mg / kg, is administered. 8. Use of D-cycloserine (DCS) in a formulation to increase exploration behavior and perceptiveness. 9. Use according to claim 8, characterized in that D-cycloserine is formulated for oral or parenteral administration, in particular for intravenous, intramuscular, subcutaneous, transdermal, transmucosal or inhalatory administration. 10. Use according to claim 8 or 9, characterized in that D-cycloserine is formulated as a nutritional supplement, aerosol, capsule, powder, solution, suspension, tablet, injection, infusion, gel, paste or transdermal patch. Use according to any one of claims 8 to 10, characterized in that the formulation comprises at least one pharmaceutically acceptable excipient, carrier and / or diluent. 12.Use according to one of claims 8 to 11, characterized in that D-cycloserine in an amount of 0.5 to 50 mg / kg, preferably 1 to 40 mg / kg, more preferably 1.5 to 30 mg / kg, in particular 2 to 20 mg / kg. SUBSEQUENT