Claims:1. A compound for the treatment and management of nicotine addiction, wherein the compound is a selective Orexin receptor antagonist.
2. The compound as claimed in claim 1, wherein the selective Orexin receptor antagonist compound is SB334867.
3. The compound as claimed in claim 1, wherein SB-334867 is 1-(2-methylbenzoxazol-6-yl)-3-[1,5] naphthyridin-4-yl urea hydrochloride.
4. The compound as claimed in claim 1, wherein nicotine dependence induced withdrawal syndrome in mice is induced by mecamylamine at a dose of 3 mg/kg in mice.
5. The compound as claimed in claim 1, wherein SB334867 dose dependently attenuates the mecamylamine precipitated withdrawal severity score, piloerection, jumping frequency, hyperalgesia and anxiety in nicotine dependent mice.
6. The compound as claimed in claim 1, wherein SB-334867 produces significant antinociceptive effect in nicotine induced hyperalgesia in nicotine dependent mice.
7. The compound as claimed in claim 1, wherein effective dosage of SB-334867 is in the range of 0.1 to 30 mg/kg, preferably in the range of 1 to 20mg/kg.
8. The compound as claimed in claim 1, wherein preferred route of administration of SB-334867 to mice is i.p.
9. The compound as claimed in claim 1, wherein withdrawal efficacy of the compound is measured by jumping behavior, tail flick latency, piloerection behavior, body tremor behavior or by the average time spent in the open arm and head dipping frequency.

Description:FIELD OF THE INVENTION
[0001] The present invention relates to the field of pharmaceuticals. Specifically, the present invention relates to a compound that attenuates nicotine dependence induced withdrawal syndrome. The present invention further relates to a compound that is for the treatment and management of nicotine addiction, wherein the compound is a selective Orexin receptor antagonist.

BACKGROUND OF THE INVENTION
[0002] Background description includes information that may be useful in understanding the present invention. It is not an admission that any of the information provided herein is prior art or relevant to the presently claimed invention, or that any publication specifically or implicitly referenced is prior art.
[0003] Nicotine addiction is serious health problem which resulting in millions of preventable deaths worldwide. Nicotine acts on nicotinic cholinergic receptors, which demonstrate diversity in subunit structure, function, and distribution within the nervous system, presumably mediating the complex actions of nicotine described in tobacco users. Tobacco use is a major worldwide health problem (Glantz & Leshner, 2000). The World Health Organization estimates that one-third of the global adult population smokes. Tobacco is addictive, and it is difficult to quit smoking. More than 80% of the attempts to quit smoking fail within a year, and those who succeed usually have tried to quit repeatedly (Zhou et al., 2009). The accumulated evidence from a wide range of studies indicates that nicotine is the major addictive component of tobacco that drives continued use despite the harmful consequences (Dani & Biasi, 2001). Pause or discontinuation of tobacco intake precipitate both somatic and effective symptoms of withdrwal this include severe craving for nicotine anxiety, loss of concentration, restlessness, decreased heart rate, decreased mood, impatience, insomnia, increased appetite and weight gain (Pergadia et al., 2014). These withdrawal symptoms produce enough axiety and discourage persons taking large amount of nicotine to practice restraining oneself from indulging in nicotine which produces drive to relapse.
[0004] Orexins/hypocretins are hypothalamic peptides involved in arousal and wakefulness, but also play a critical role in drug addiction and reward-related behaviours (Walker & Lawrence, 2017). There are two receptors system for orexin, termed OxR1 and OxR2. Both receptors couple with G proteins; OxR1 couples exclusively with Gq subunits while OxR2 couples to both Gq and Gi/o (Urbanska et al., 2012). Orexin neurons send dense fiber projections throughout the brain including cerebral cortex, hippocampus, thalamus, midbrain and spinal cord. Furthermore, OxR1 and OxR2 are widely distributed throughout the CNS (Shan et al., 2015). Orexin has been implicated in feeding behavior, in keeping with the location of these cells in the area of lateral hypothalamus (LH). Much attention has been focused recently on the role of orexin in drug reward (Nishino et al., 2000). The hypothalamus is a prominent central site of nicotine action that may mediate the effects of this drug on endocrine, circadian, reward and ingestive parameters. Furthermore, chronic nicotine treatment upregulates expression of orexin peptides and receptors. Hypothalamic orexin neurons are important regulators of state-dependent behavior, arousal and feeding but also play a critical role in drug addiction and reward-related behaviors in nicotine dependent rodents. Pharmacological modulation of the orexin system can affect behavioral responses seen during nicotine withdrawal syndrome in rodents.
[0005] The current drugs used in the clinic to treat nicotine dependence have been shown to just contain the symptomatology of the withdrawal syndrome rather than affecting the pathophysiological course of the disease and various adverse effects linked with them (Marlatt & Donovan, 2005). Therefore, research is being carried out in order to find out pharmacological approaches that might ameliorate the problem of nicotine dependence. The initial pharmacogenetics studies of pharmaco-therapies that have recently been approved by the united states food and drug adminstration of following for the treatment of nicotine dependence nicotine replacement therapy (using nicotine gum, nicotine nasal spray, and transdermal nicotine) and bupropion (Watkins et al., 2000).
[0006] Numerous studies have shown that orexin plays important roles in drug seeking in animal models of addiction. However, the exact nature of these roles has remained elusive. This is most likely due to a complex involvement of orexin in several aspects of drug seeking, such as aversive as well as appetitive motivation, interactions with Pavlovian and/or instrumental learning processes, and hedonic states induced by drugs. Therefore, the neural circuitry in which orexin is embedded becomes very important for understanding the role of orexin in drug seeking and reward. (Minami et al., 2007).
[0007] Thus there is an urgent need to develop new compounds that can attenuate nicotine dependence induced withdrawal syndrome, can be used to tackle the problem of nicotine addiction and can overcome deficiencies associated with the known arts.

OBJECTS OF THE INVENTION
[0008] An object of the present invention is to provide a compound that can attenuate nicotine dependence induced withdrawal syndrome.
[0009] Yet another object of the present invention is to provide a compound that is useful for treatment and management of nicotine addiction.
[0010] Another object of the present invention is to provide a compound that provides dose dependent attenuation in the withdrawal syndrome.
[0011] Still another object of the present invention is to provide a compound that provides in-vivo quantitative results to attenuate nicotine dependence induced withdrawal syndrome in mice.
[0012] Another object of the present invention is to provide compounds that are safe to use and exhibit no severe side effects.
[0013] The other objects and preferred embodiments and advantages of the present invention will become more apparent from the following description of the present invention when read in conjunction with the accompanying examples and figures, which are not intended to limit scope of the present invention in any manner.

SUMMARY OF THE INVENTION
[0014] This summary is provided to introduce a selection of concepts in a simplified form that are further described below in Detailed Description section. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used as an aid in determining the scope of the claimed subject matter.
[0015] The present invention relates to the field of pharmaceuticals. Specifically, the present invention relates to a compound that attenuates nicotine dependence induced withdrawal syndrome.
[0016] The present invention further relates to a compound for the treatment and management of nicotine addiction, wherein the compound is a selective Orexin receptor antagonist.
[0017] In one aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein the selective Orexin receptor antagonist compound is SB334867.
[0018] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein the compound SB-334867 is 1-(2-methylbenzoxazol-6-yl)-3-[1,5] naphthyridin-4-yl urea hydrochloride.
[0019] In yet another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB334867 acts as a selective OX1r antagonist.
[0020] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein nicotine dependence induced withdrawal syndrome in mice is induced by mecamylamine at a dose of 3 mg/kg in mice.
[0021] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB334867 dose dependently attenuates the mecamylamine precipitated withdrawal severity score, piloerection, jumping frequency, hyperalgesia and anxiety in nicotine dependent mice.
[0022] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB-334867 produces significant antinociceptive effect in nicotine induced hyperalgesia in nicotine dependent mice.
[0023] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein effective dosage of SB-334867 is in the range of 0.1 to 30 mg/kg.
[0024] In another aspect, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein withdrawal efficacy of the compound is measured by jumping behavior, tail flick latency, piloerection behavior, body tremor behavior or by the average time spent in the open arm and head dipping frequency.
[0025] Other aspects of the invention will be set forth in the description which follows, and in part will be apparent from the description, or may be learnt by the practice of the invention.

BRIEF DESCRIPTION OF DRAWINGS THE INVENTION
[0026] The following drawings form part of the present specification and are included to further illustrate aspects of the present disclosure. The disclosure may be better understood by reference to the drawings in combination with the detailed description of the specific embodiments presented herein.
Figure 1: Effect of treatment(s) on mecamylamine induced increase in jumping frequency in nicotine dependent/ vehicle treated mice
Figure 2: Effect of treatment(s) on mecamylamine induced increase in withdrawal severity score in nicotine dependent/ vehicle treated mice
Figure 3: Effect of treatment(s) on mecamylamine induced hyperalgesia in nicotine dependent/ vehicle treated mice
Figure 4: Effect of treatment(s) on mecamylamine induced piloerection in nicotine dependent/ vehicle treated mice
Figure 5: Effect of treatment(s) on mecamylamine induced Body tremor in nicotine dependent/ vehicle treated mice
Figure 6: Effect of treatment(s) on mecamylamine induced anxiety like behaviour in nicotine dependent/ vehicle treated mice in terms of time spent in open arm in elevated plus maze test
Figure 7: Effect of treatment(s) on mecamylamine induced anxiety like behaviour in nicotine dependent/ vehicle treated mice in terms of time spent in Close arm in elevated plus maze test
Figure 8: Effect of treatment(s) on mecamylamine induced anxiety like behaviour in nicotine dependent/ vehicle treated mice in terms of time spent in centre platform in elevated plus maze test
Figure 9: Effect of treatment(s) on mecamylamine induced anxietylike behaviour in nicotine dependent/ vehicle treated mice in terms of Frequency of head-dipping & stretched attend posture in elevated plus maze test

DETAILED DESCRIPTION
[0027] The following is a detailed description of embodiments of the disclosure. The embodiments are in such detail as to clearly communicate the disclosure. However, the amount of detail offered is not intended to limit the anticipated variations of embodiments; on the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the present disclosure as defined by the appended claims.
[0028] All publications herein are incorporated by reference to the same extent as if each individual publication or patent application were specifically and individually indicated to be incorporated by reference. Where a definition or use of a term in an incorporated reference is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.
[0029] Reference throughout this specification to “one embodiment” or “an embodiment” means that a particular feature, structure or characteristic described in connection with the embodiment is included in at least one embodiment. Thus, the appearances of the phrases “in one embodiment” or “in an embodiment” in various places throughout this specification are not necessarily all referring to the same embodiment. Furthermore, the particular features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.
[0030] In some embodiments, the numbers expressing quantities of ingredients, properties such as concentration, reaction conditions, and so forth, used to describe and claim certain embodiments of the invention are to be understood as being modified in some instances by the term “about.” Accordingly, in some embodiments, the numerical parameters set forth in the written description and attached claims are approximations that can vary depending upon the desired properties sought to be obtained by a particular embodiment. In some embodiments, the numerical parameters should be construed in light of the number of reported significant digits and by applying ordinary rounding techniques. Notwithstanding that the numerical ranges and parameters setting forth the broad scope of some embodiments of the invention are approximations, the numerical values set forth in the specific examples are reported as precisely as practicable. The numerical values presented in some embodiments of the invention may contain certain errors necessarily resulting from the standard deviation found in their respective testing measurements.
[0031] As used in the description herein and throughout the claims that follow, the meaning of “a,” “an,” and “the” includes plural reference unless the context clearly dictates otherwise. Also, as used in the description herein, the meaning of “in” includes “in” and “on” unless the context clearly dictates otherwise.
[0032] Unless the context requires otherwise, throughout the specification which follow, the word “comprise” and variations thereof, such as, “comprises” and “comprising” are to be construed in an open, inclusive sense that is as “including, but not limited to.”
[0033] The recitation of ranges of values herein is merely intended to serve as a shorthand method of referring individually to each separate value falling within the range. Unless otherwise indicated herein, each individual value is incorporated into the specification as if it were individually recited herein. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples, or exemplary language (e.g. “such as”) provided with respect to certain embodiments herein is intended merely to better illuminate the invention and does not pose a limitation on the scope of the invention otherwise claimed. No language in the specification should be construed as indicating any non-claimed element essential to the practice of the invention.
[0034] Groupings of alternative elements or embodiments of the invention disclosed herein are not to be construed as limitations. Each group member can be referred to and claimed individually or in any combination with other members of the group or other elements found herein. One or more members of a group can be included in, or deleted from, a group for reasons of convenience and/or patentability. When any such inclusion or deletion occurs, the specification is herein deemed to contain the group as modified thus fulfilling the written description of all Markush groups used in the appended claims.
[0035] The description that follows, and the embodiments described therein, is provided by way of illustration of an example, or examples, of particular embodiments of the principles and aspects of the present disclosure. These examples are provided for the purposes of explanation, and not of limitation, of those principles and of the disclosure.
[0036] It should also be appreciated that the present disclosure can be implemented in numerous ways, including as a system, a method or a device. In this specification, these implementations, or any other form that the invention may take, may be referred to as processes. In general, the order of the steps of the disclosed processes may be altered within the scope of the invention.
[0037] The headings and abstract of the invention provided herein are for convenience only and do not interpret the scope or meaning of the embodiments.
[0038] The following discussion provides many example embodiments of the inventive subject matter. Although each embodiment represents a single combination of inventive elements, the inventive subject matter is considered to include all possible combinations of the disclosed elements. Thus if one embodiment comprises elements A, B, and C, and a second embodiment comprises elements B and D, then the inventive subject matter is also considered to include other remaining combinations of A, B, C, or D, even if not explicitly disclosed.
[0039] Various terms as used herein are shown below. To the extent a term used in a claim is not defined below, it should be given the broadest definition persons in the pertinent art have given that term as reflected in printed publications and issued patents at the time of filing..
[0040] The present invention relates to the field of pharmaceuticals. Specifically, the present invention relates to a compound that attenuates nicotine dependence induced withdrawal syndrome.
[0041] The present invention further relates to a compound for the treatment and management of nicotine addiction, wherein the compound is a selective Orexin receptor antagonist.
[0042] In one embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein the selective Orexin receptor antagonist compound is SB334867.
[0043] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein the compound SB-334867 is 1-(2-methylbenzoxazol-6-yl)-3-[1,5] naphthyridin-4-yl urea hydrochloride.
[0044] In yet another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB334867 acts as a selective OX1r antagonist.
[0045] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein nicotine dependence induced withdrawal syndrome in mice is induced by mecamylamine at a dose of 3 mg/kg in mice.
[0046] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB334867 dose dependently attenuates the mecamylamine precipitated withdrawal severity score, piloerection, jumping frequency, hyperalgesia and anxiety in nicotine dependent mice.
[0047] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein SB-334867 produces significant antinociceptive effect in nicotine induced hyperalgesia in nicotine dependent mice.
[0048] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein effective dosage of SB-334867 is in the range of 0.1 to 30 mg/kg.
[0049] In another preferred embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein effective dosage of SB-334867 is in the range of 1 to 20 mg/kg.
[0050] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein preferred route of administration of SB-334867 to mice is i.p.
[0051] In another embodiment, the present invention relates to a compound for the treatment and management of nicotine addiction, wherein withdrawal efficacy of the compound is measured by jumping behavior, tail flick latency, piloerection behavior, body tremor behavior or by the average time spent in the open arm and head dipping frequency.
[0052] While the foregoing describes various embodiments of the disclosure, other and further embodiments of the disclosure may be devised without departing from the basic scope thereof. The scope of the invention is determined by the claims that follow. The invention is not limited to the described embodiments, versions or examples, which are included to enable a person having ordinary skill in the art to make and use the invention when combined with information and knowledge available to the person having ordinary skill in the art.
[0053] The present invention is further explained in the form of following examples. However, it is to be understood that the following examples are merely illustrative and are not to be taken as limitations upon the scope of the invention.
[0054] Materials and Methods: Swiss albino male mice weighing 25±2g, obtained from Central Research Institute, Kasauli, India and maintained on standard laboratory diet (Kisan Feeds Ltd., Mumbai, India), having free access to tap water were employed in the present study. They were housed in the animal house and were exposed to a regular 12 hr cycle of light and dark. The experiments were conducted in a semi-sound proof laboratory. The observer was blind to the treatment group assignment. The experimental protocol was approved by the institutional animal ethical committee and care of the animals was done as per the guidelines of Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA), Ministry of Environment and Forests, Government of India (Reg. No. 1181/PO/ReBi/S/08/CPCSEA).
[0055] Drugs and chemicals: Mecamylamine hydrochloride, SB-334867 and nicotine tartrate (procured from Sigma-Aldrich Chemicals Pvt. Ltd., St. Louis, USA), were dissolved / diluted in normal saline and DMSO. The chemicals used were of analytical grade and all drug solutions were freshly prepared before use.
[0056] Example 1: Induction of nicotine withdrawal syndrome in mice
[0057] Sub-acute administration of nicotine followed by a single injection of mecamylamine was used to induce nicotine withdrawal in mice (Damaj et al., 2003; Biala and Weglinska, 2005; Singh et al., 2013). Nicotine dependence was induced by repeated s.c. injections, four times daily, at an interval of 4 h starting at 06:00 a.m, for 7 days (2.5 mg/kg, s.c). The control groups were treated with saline following the same schedule. On the test day (day 8), mice received one nicotine injection in the morning. In attempt to precipitate nicotine abstinence mice were given the nicotine receptor antagonist mecamylamine (3 mg kg-1, i.p.), 1 h after the last nicotine injection. The somatic signs of withdrawal were evaluated for 30 min, immediately after mecamylamine administration. The observations were made in a transparent perspex observation chamber with dimensions of 30 cm X 30 cm X 30 cm.
[0058] Example 2: Quantitative Assessment of withdrawal syndrome
[0059] Assessment of Nicotine withdrawal syndrome in terms of jumping frequency: Stereotyped jumps precipitated by nicotine receptor antagonist mecamylamine (3 mg kg-1, i.p.), 1 h after the last nicotine injection, has been considered as a predominant sign for quantification of Nicotine withdrawal syndrome in mice (Biala and Weglinska, 2005; Singh et al., 2013 a, b). Jumping frequency observed in a period of 30 min was used as a quantitative symptom of Nicotine withdrawal immediately after mecamylamine administration.
[0060] Measurement of the effect of drug treatment(s) on nicotine-induced hyperalgesia using tail flick test: Sub-chronic administration of nicotine is documented to cause the induction of hyperalgesia which has been quantified in terms of tail flick test (Kiguchi et al. 2008; Jackson et al. 2010). Therefore, nociceptive threshold was measured by the tail flick test in mice (D'Amour and Smith 1941; Singh et al., 2013 a, b). A cut off latency time was fixed at 10 sec. Thus, tail flick latency was observed five minutes later, after mecamylamine administration.
[0061] Assessment of anxiety like behaviour using elevated plus maze test: Anxiety like behaviour was monitored in mice using an elevated plus maze test (Navarro et al., 2006; Singh et al., 2013 a, b). The mouse is placed in the center of the maze, facing one of the enclosed arms. During a 5 min test period the following measures were taken: the number of entries into and time spent in the open and enclosed. Likewise, different ethological measures were also quantified: (a) Stretched attend posture (SAP): a body posture in which the mouse stretches forward and then retracts to its original position without moving the feet, and (b) Head-dipping (HD): movement of the head over the side of the maze and down towards the floor. The elevated plus maze test was performed 30 minutes after the administration of mecamylamine on day 8 of the nicotine dependence procedure to assess the level of anxiety like behavior in mice before and after the treatment schedule is over.
[0062] Assessment of nicotine withdrawal syndrome in terms of piloerection frequency: Piloerection frequency observations were made for a period of 30 min to quantitate the severity of the experimental withdrawal phenomenon immediately after mecamylamine administration. This parameter has been noted to be indicative of the intensity ofwithdrawal syndrome as also reported earlier (Damaj et al. 2003; Biala and Weglinska 2005).
[0063] Assessment of nicotine withdrawal syndrome in terms of body tremor frequency: Body tremor frequency observations were made for a period of 30 min to quantitate the severity of the experimental withdrawal phenomenon immediately after mecamylamine administration. These parameters have been noted to be indicative of the intensity of withdrawal syndrome as also reported earlier (Damaj et al. 2003; Biala and Weglinska 2005).
[0064] Example 3: Experimental protocol
[0065] Seven groups were employed in the present study, with each group comprising of 08 animals out of which half were males and half females.
[0066] Group I (Vehicle-vehicle control): Vehicle (Saline, 10 ml/kg, i.p.) for Nicotine was administered four times daily for a period of seven days. Vehicle (10% DMSO in water, 10 ml/kg, i.p.) for SB-334867 was simultaneously injected once daily for the same period of seven days. Vehicle (10 ml/kg, i.p.) for Mecamylamine was then injected on the morning of day 8, 1 hr after administering vehicle (Saline, 10 ml/kg, i.p.) for nicotine.
[0067] Group II (Vehicle-mecamylamine control): Vehicle (Saline, 10 ml/kg, i.p.) for Nicotine was administered four times daily for a period of seven days. Vehicle (10% DMSO in water, 10 ml/kg, i.p.) for SB-334867 was simultaneously injected once daily for the same period of seven days. Mecamylamine (3 mg/kg, i.p.) was then injected on the morning of day 8, 1 hr after administering vehicle (Saline, 10 ml/kg, i.p.) for nicotine.
[0068] Group III (Nicotine–Mecamylamine control): Nicotine (2.5 mg/ kg, s.c.) was administered four times daily for a period of seven days. Vehicle (10% DMSO in water, 10 ml/kg, i.p.) for SB-334867 was simultaneously injected once daily for the same period of seven days. Mecamylamine (3 mg/kg, i.p.) was then injected on the morning of day 8, 1 hr. after administering Nicotine (2.5 mg/ kg, s.c.).
[0069] Group IV (SB-334867 treatment + Vehicle- Mecamylamine control): Vehicle (Saline, 10 ml/kg, i.p.) for nicotine was administered four times daily for a period of 7 days. SB-334867 (20 mg/kg, i.p.) was simultaneously injected once daily for the same period of 7 days. Mecamylamine (8 mg/kg, i.p.) was then injected on the morning of day 8, 1 hr. after administering vehicle (Saline, 10 ml/kg, i.p.) for Nicotine.
[0070] Group V-VII (SB-334867 treatment + nicotine-mecamylamine): nicotine (2.5 mg/ kg, s.c.) was administered four times daily for a period of seven days. SB-334867 (at a dose level of 1, 10 and 20 mg/kg/d, i.p. for groups number V, VI, and VII, respectively) was simultaneously injected once daily for the same period of seven days. Mecamylamine (3 mg/kg, i.p.) was then injected on the morning of day 8, 1 hr. after administering nicotine (2.5 mg/ kg, s.c.).
[0071] Example 4:Statistical analysis and Results
[0072] All the results were expressed as mean ± standard error of mean (S.E.M.). Data of the results was analyzed using ANOVA followed by post-hoc comparison using Sheffe’s multiple range test. For elevated plus maze, the numbers of entries and time (in seconds) spent in both arms was compared in saline- and nicotine-treated mice. A value of P<0.05 was considered to be statistically significant. The statistical analysis was done using Sigma Stat 6.0 software.
[0073] Effect of SB-334867 treatments on mecamylamine induced alteration in jumping behavior in nicotine dependent mice
[0074] Nicotine (2.5 mg/ kg, s.c.) was administered four times daily for a period of seven days. On the test day (day 8), mice received one nicotine injection in the morning. In attempt to precipitate nicotine abstinence mice were given the nicotine receptor antagonist mecamylamine (3 mg kg-1, i.p.), 1h after the last nicotine injection precipitated withdrawal syndrome in mice as reflected by a significant increase (p<0.01) in stereotyped jumping behavior in nicotine/ mecamylamine group, when compared to that of the vehicle treated control group. Administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.01 each) and dose dependently attenuated mecamylamine induced withdrawal syndrome in nicotine dependent mice, when measured in terms of stereotyped jumping behavior (Figure 1).
[0075] Effect of SB-334867 treatments on mecamylamine induced alteration in withdrawal severity score in nicotine dependent mice
[0076] Administration of four doses of nicotine (2.5 mg/kg, s.c) for a period of 7 days, followed by a single injection of mecamylamine (3mg/kg, i.p.) precipitated withdrawal syndrome in mice as reflected by a significant increase (p<0.01) in the composite withdrawal severity score (Rehni et al., 2012; Singh et al., 2013 a, b) measured in terms of a concomitant rise in the severity of grooming, scratching, chewing, fore paw tremor, wet dog shake, cage scratching, head nodding, paw licking, particularly in the nicotine-mecamylamine group, when compared to that of the vehicle treated control groups. Administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.01 each) and dose dependently attenuated nicotine-mecamylamine induced withdrawal syndrome in mice, when measured in the terms of the total withdrawal severity score accompanied by a marked decrease in severity of the observed behavioral criteria viz., grooming, scratching, chewing, fore paw tremor, wet dog shake, cage scratching, head nodding, paw licking (Figure 2).
[0077] Effect of SB-334867treatment(s) on nicotine-induced hyperalgesia using tail flick test
[0078] After chronic intermittent administration of nicotine (2.5 mg kg-1, 7 days, four injections/day) produced significant hyperalgesia, as indicated by decreased tail-flicklatencies (P <0.05), when compared to vehicle control group. Moreover, pair-wise comparisons confirmed that administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.05 each) and dose dependently attenuated nicotine-mecamylamine induced hyperalgesia measured in terms reversal of withdrawal induced decrease in the tail flick latency (Figure 3).
[0079] Effect of SB-334867treatment(s) on piloerection behaviour in nicotine dependent mice
[0080] Statistical analysis showed that there was a main effect of group in ANOVA when applied to piloerection frequency data. Nicotine (2.5 mg/ kg, s.c.) was administered four times daily for a period of seven days. On the test day (day 8), mice received one nicotine injection in the morning. In attempt to precipitate nicotine abstinence mice were given the nicotine receptor antagonist mecamylamine (3 mg/kg, i.p.), 1h after the last nicotine injection precipitated withdrawal syndrome in mice as reflected by a significant increase (p<0.05) in stereotyped piloerection behaviour in nicotine/ mecamylamine group, when compared to that of the vehicle treated control group. Further, pair-wise comparisons confirmed that administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.01 each) and dose dependently attenuated mecamylamine induced withdrawal syndrome in nicotine dependent mice, when measured in terms of stereotyped piloerection behavior (Figure 4).
[0081] Effect of SB-334867treatment(s) on body tremor in nicotine dependent mice
[0082] Statistical analysis showed that there was a main effect of group in ANOVA when applied to body tremor frequency data. Nicotine (2.5 mg/ kg, s.c.) was administered four times daily for a period of seven days. On the test day (day 8), mice received one nicotine injection in the morning. In attempt to precipitate nicotine abstinence mice were given the nicotine receptor antagonist mecamylamine (3 mg/kg, i.p.), 1 h after the last nicotine injection precipitated withdrawal syndrome in mice as reflected by a significant increase (p<0.05) in stereotyped tremor behaviour in nicotine/ mecamylamine group, when compared to that of the vehicle treated control group. Further, pair-wise comparisons confirmed that administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.01 each) and dose dependently attenuated mecamylamine induced withdrawal syndrome in nicotine dependent mice, when measured in terms of stereotyped body tremor behavior (Figure 5).
[0083] Effect of SB-334867 treatments on mecamylamine induced anxiety like behaviour in nicotine dependent mice
[0084] Administration of four doses of nicotine (2.5 mg/kg, s.c) for a period of 7 days, followed by a single injection of mecamylamine (3mg/kg, i.p.) precipitated withdrawal syndrome in mice as reflected by a significant increase (p<0.01) in anxiety like behaviour as measured in terms of average time spent in the open arm and head dipping frequency (P<0.01) particularly in the nicotine-mecamylamine group, when compared to that of the vehicle treated control groups. Administration of SB-334867 (1, 10 & 20 mg/kg, i.p.) significantly (p<0.01 each) and dose dependently attenuated nicotine-mecamylamine induced anxiety like behaviour as measured in terms of the average time spent in the open arm, close arm, centre platform and head dipping frequency (Figure 6, 7, 8 & 9).
[0085] The foregoing examples are merely illustrative and are not to be taken as limitations upon the scope of the invention. Various changes and modifications to the disclosed embodiments will be apparent to those skilled in the art. Such changes and modifications may be made without departing from the scope of the invention.