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Effects of chronic nicotine administration on body weight, food intake and nitric oxide concentration in female and male rats
Pathophysiology 21 (2014) 185–190
Effects of chronic nicotine administration on body weight, food intake and nitric oxide concentration in female and male rats Omamuyovwi Meashack Ijomone a,b,∗ , Olayemi Kafilat Olaibi a , Polycarp Umunna Nwoha a b
a Department of Anatomy and Cell Biology, Obafemi Awolowo University, Ile-Ife, Osun, Nigeria Department of Human Anatomy, Cross River University of Technology, Okuku, Cross River, Nigeria
Abstract Nicotine is readily consumed through cigarettes; however it is also easily consumed through the various forms of non-prescription nicotine replacement therapy. It has been shown to possess potential therapeutic value for the management of neurologic and neurodegenerative diseases in the last decade. Hence, this study examined the effects of chronic subcutaneous nicotine administration on food intake and body weight as well as on nitric oxide concentrations and total antioxidant capacity in female and male rats. Nicotine was administered to rats via subcutaneous injections at doses of 0.25, 2 and 4 mg/kg body weight for 28 days. Control groups received normal saline; the vehicle for nicotine. Food intake by each group was monitored daily and body weight of the animals was measured twice weekly. At the end of drug administration, blood was obtained from each animal via cardiac puncture for biochemical determination of serum total antioxidant capacity (TAC) and nitric (NO) concentrations using standard assay kits. Results show significant loss (p < 0.05) of body weight in all nicotine treated female rats. In contrast, male rats showed weight gain, though this was significantly lower (p < 0.001) in nicotine treated groups compared to control. Nicotine significantly reduced (p < 0.001) food consumed in both female and male rats; however dose related changes were observed in only male rats. No significant difference was observed in TAC following nicotine treatments for both female and male rats. Furthermore, only males exhibited changes in NO concentrations following nicotine treatment, as it significantly increased (p < 0.01) NO concentrations in all male treated groups. In conclusion, this study has shown that modulation of body weight, food consumption and nitric oxide formation by nicotine is sexually dimorphic. Also, the study suggests that nicotine modulation of food intake and body weight and its modulation of NO may be independent of each other. © 2014 Elsevier Ireland Ltd. All rights reserved.
Keywords: Nicotine; Body weight; Food intake; Nitric oxide; Sexual differences
1. Introduction Nicotine the natural alkaloid present in Nicotiana tabacum is a readily available substance. It is the main component of cigarettes. However it is also easily consumed through the various forms of non-prescription nicotine replacement therapy such as nicotine dermal patches and nicotine chewing gums [1]. Concerns continually exist on the global effects of nicotine exposure in the body; nevertheless it is well known ∗ Corresponding author at: Department of Anatomy and Cell Biology, Obafemi Awolowo University, Ile-Ife, Osun, Nigeria. Tel.: +234 7031354971. E-mail addresses: [email protected], [email protected] (O.M. Ijomone).
http://dx.doi.org/10.1016/j.pathophys.2014.08.003 0928-4680/© 2014 Elsevier Ireland Ltd. All rights reserved.
that many of the detrimental effects associated with smoking are mostly due to the combination of many other constituents of cigarette smoke, and not nicotine in itself [2]. Though nicotine is the major reason smoking can be addictive, because of its stimulation of the reward-based system within the brain [3], the toxic nature of the many other constituents of cigarettes and their reactions, is largely responsible for serious illness such as coronary heart disease, emphysema and varying forms of cancer caused by smoking [2]. Nevertheless, when administered alone especially by transdermal patches or intravenously, nicotine is known to produce only little or no health problems [2]. Reports indicate an inverse relationship between smoking and body weight, which appears to be mediated by nicotine [4–6]. It has been suggested that nicotine modifies body
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weight via suppression of food intake possible by modulating levels of neuroregulatory substances known to activate or suppress feeding [5,7]. Nicotine has also been seen to have regulatory actions on nitric oxide (NO) synthesis. NO, a neural messenger in the central and peripheral nervous systems is generated from l-arginine by nitric oxide synthase (NOS), following the activation of NMDA receptors [8,9]. In neurons, NO synthesis occurs in an intricate sequence of events that involves stimulation of NMDA-subtype glutamate recetors and influx of Ca2+ into the cytoplasm which upon binding to calmodulin activates neuronal NOS (nNOS) [10]. Stimulation of nAChRs by nicotine is known to activate NMDA receptors and increase intracellular Ca2+ , thus resulting in formation of NO [10,11]. On the other hand, NO has been shown to modulate feeding behaviours. Inhibition of NO synthesis reduces food intake and body weight [12,13]. l-Arginine from which NO is generated results in increased food intake and antagonizes leptin reduction in food intake and body weight [14,15]. Nicotine has been the subject of potential therapeutic value for the management of neurologic and neurodegenerative diseases in the last decade [16,17]. In this study, we have examined the effects of chronic subcutaneous nicotine administration on food intake and body weight as well as on nitric oxide concentrations and total antioxidant capacity. We have used rats of both sexes with a view of appraising sexual dimorphic responses. 2. Materials and methods 2.1. Animal management and treatment Female and male adult albino strain Wistar rats (150–200 g) were used for this study. Animals were housed in clean plastic cages in a clean environment of 12 h day/light cycle, at room temperature. Animals in all groups were allowed access to standard laboratory rat chow and water ad libitum. All experimental protocols were in strict accordance with the guidelines for animal research, as detailed in the NIH Guidelines for the Care and Use of Laboratory Animals (National Academy of Sciences and National Institutes of Health Publications, 2011) and approved by local Institutional Research Committee. A total of 40 rats (20 females and 20 males) were randomly grouped into 4 groups of 5 rats each; a control group and 3 treated groups each for both female and male rats. Treated groups were administered nicotine via subcutaneous injections at doses of 0.25, 2 and 4 mg/kg body weight for 28 days. Control groups received normal saline – vehicle for nicotine. Nicotine was obtained in free base form as (−)nicotine also called (−)-1-methyl-2-(3-pyridyl)pyrrolidine from Sigma Chemicals, USA. Food intake by each group was monitored daily. Weekly or total food consumed was expressed as mean of daily consumptions for each week or throughout the 28-day period of administration. Body weight of the animals was measured twice weekly. Weight gain/loss
was determined by subtracting the body weight at the beginning of administration (initial body weight) from the body weight at end of administration (final body weight). At the end of the administration, animals were euthanized and blood was obtained from each animal via cardiac puncture for biochemical determination of serum TAC and NO concentrations using standard assay kits. 2.2. Biochemical determination of TAC and NO concentrations TAC was determined using the QuantiChromTM Antioxidant Assay Kit (BioAssays Systems, USA). This BioAssay Systems’ improved assay measures total antioxidant capacity in which Cu2+ is reduced by antioxidant to Cu+ . The resulting Cu+ specifically forms a coloured complex with a dye reagent. NO was determined using the QuantiChromTM Nitric Oxide Assay Kit (Bioassays Systems, USA) for the Colorimetric determination of total nitrite and based on the enzymatic conversion of nitrate to nitrite by nitrate reductase. BioAssay Systems’ QuantiChromTM Nitric Oxide Assay Kit is designed to accurately measure NO production following reduction of nitrate to nitrite using improved Griess method. These biochemical studies were carried out following protocols in manufacturer’s guidelines. 2.3. Statistical analysis Data obtained were analyzed using One-way ANOVA, followed by Student Newman–Keuls (SNK) for post-tests. GraphPad Prism 5 (Version 5.03, GraphPad Software, USA.) was the statistical package used for data analysis. Student’s t-test was employed for analysis of between group differences, when appropriate. Statistical significance was set at p < 0.05. 3. Results 3.1. Food intake and body weight The present study observed a marked decrease in weekly body weight in all nicotine treated female groups. Consequently, there was a significant loss of weight in all nicotine treated female groups compared to control (p < 0.05) (Fig. 1). In contrast, male rats showed a steady rise in weekly body weight for all nicotine treated groups, however weight gain was significantly lowered in all nicotine treated male groups compared to control (p < 0.001) (Fig. 1). No dose related changes in body weight gain/loss was observed for both female and male nicotine treated groups. Total food consumed by female rats was significantly reduced in all nicotine treatment groups compared to control (p < 0.001) (Fig. 2). However, no dose related changes were observed. Contrastingly, male rats showed dose related changes following nicotine treatment. Total food consumed by male rats was significantly reduced in all nicotine
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Fig. 1. Line graph of weekly body weight and weight gain/loss of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. *p < 0.05, **p < 0.01, ***p < 0.001. α, β, and δ – significant difference compared to control, 0.25 mg/kg and between 2 mg/kg and 4 mg/kg respectively. One way ANOVA followed by SNK for post-tests.
treatment groups compared to control (p < 0.001). However, 2 mg/kg (72.93 ± 1.95) and 4 mg/kg (77.18 ± 2.28) treatments significantly reduced food intake compared to 0.25 mg/kg (82.93 ± 1.32) nicotine treatment (p < 0.001) (Fig. 2). 3.2. TAC and NO concentrations No significant difference was observed in TAC following nicotine treatments for both male and female rats. Also, no significant difference was observed in NO concentrations of nicotine treated female rats compared to control. In contrast, NO concentrations were significantly increased in all nicotine treatment groups compared to control in male rats (p < 0.01). However no dose related changes were observed (Fig. 3).
4. Discussion The present study showed that chronic subcutaneous administration of nicotine produced sexual dimorphic response in body weight and food consumed. Nicotine resulted in weight loss in females but produced only a slight weight gain in males. Also, nicotine produced dose dependent reduction in total food consumed in males; however, reduction in total food consumed was not dose dependent in females. Reduction in body weight and food intake is a
more consistent effect associated with nicotine administration and tobacco smoking. Numerous studies in both human and animal studies have demonstrated that nicotine and tobacco suppresses body weight and food intake [4–6,18–20]. Chronic nicotine administration has been shown to decrease body weight and food intake [6,7], supporting data from the present study. It has been argued that changes in body weight induced by nicotine results from changes in food intake [6,7], as also supported by this study. In addition, nicotine has been shown to increase lipolysis as well as increase fat utilization. Hence, it has been suggested that nicotine induced changes in whole-body metabolism may also contribute to its weight modulating effects [6]. However, this study has revealed sexual difference in nicotine modulation of body weight and food consumption. Fat deposits are greater in females than males, hence it is possible that nicotine produces a higher fat utilization in females than males and this may contribute to its greater reduction of body weight in females. Also, it is likely that sexual difference exists in nicotine modulation of neuroregulatory substances that suppress and activate feeding [5]. The present study revealed no significant changes in TAC following nicotine treatment compared to control in both females and males. This indicates a lack of effect on antioxidants and possibly in oxidative stress. Evidence has shown that nicotine has both pro-oxidants and antioxidants effects [21]. In vitro studies using Chinese hamsters’ ovary cells
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Fig. 2. Weekly and total food intake of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. *p < 0.05, **p < 0.01, ***p < 0.001. α, β, and δ – significant difference compared to control, 0.25 mg/kg and between 2 mg/kg and 4 mg/kg respectively. One way ANOVA followed by SNK for post-tests.
showed than nicotine induced oxidative stress and resulted in generation of ROS and hydrogen peroxide [22,23]. However, another study demonstrated that nicotine did not show antioxidant properties nor altered indicators of oxidative in vivo and in vitro [24]. Nicotine has also been demonstrated to possess antioxidant properties in experimentally induced neurodegenerative diseases such as Parkinson’s, Alzheimer’s and Huntington’s diseases [25–27]. This has prompted suggestion that its neuroprotective mechanisms may also include antioxidant actions [28]. Dose related effects have also been linked to nicotine dual-actions on oxidative stress. Generally, high doses may stimulate oxidative stress, while low doses may act as antioxidants [29]. The present findings suggest neither antioxidant effect nor induction of oxidative stress. This may not be unconnected to the recent evidence that adult rats are less sensitive to nicotine induced oxidative stress, compared to young or aged ones [30]. The present study also revealed significant increase in serum NO concentrations in all nicotine treated groups compared to control in males only but not in females. Acute or chronically administered nicotine has been demonstrated to increase NO formation in nervous system as well as in systemic organs [9,11]. Studies have reported that NO is important for synaptic plasticity and long-term potentiation (LTP) as well various cognitive and non-cognitive effects [8].
Nicotine is known to improve cognitive processes such as rapid information processing, attention, working memory and long term memory [31,32]. Increasing NO concentrations has been shown as a mechanism by which nicotine improves cognitive functions [8]. However, it is likely that this modulation of NO by nicotine and its resultant benefits may be seen only in males but not females. Also, in the present study, increased NO concentrations in male rats did not affect nicotine reduction of food intake and body weight. This is quite interesting considering that increase in NO synthesis causes increased food intake and body weight [12,14]. Hence, we suggest that nicotine modulation of food intake and body weight may be independent of NO pathways. In summary, there is growing evidence that sexually dimorphic responses are associated with nicotine exposure. These differences are important to consider in developing better treatments for smoking cessation as well as developing forms of nicotine therapy in treatment of neurodegenerative and neurologic disorders. In fact, studies have shown that women find it more difficult to quit smoking than men, and also are less likely to benefit from the use of nicotine replacement therapy in smoking cessation programmes (Perkins et al. [33,34]). Our results support this sexually dimorphic response of nicotine in respect to modulation of body weight, food consumption and nitric oxide formation. One hypothesis
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Fig. 3. TAC and NO concentrations of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. **p < 0.01. α – significant difference compared to control. One way ANOVA followed by SNK for post-tests.
to explain the difficulty to stop smoking in women is the greater perceived benefits of weight control due to smoking in women than men [35]. It is thus interesting that the present study does indeed show weight loss following nicotine administration in female rats but not in males. Also, the study suggests that nicotine modulation of food intake and body weight and its modulation of NO may be independent of one other. References [1] I. Mahar, R.C. Bagot, M.A. Davoli, S. Miksys, R.F. Tyndale, C. Walker, M. Maheu, S. Huang, T.P. Wong, N. Mechawar, Developmental hippocampal neuroplasticity in a model of nicotine replacement therapy during pregnancy and breastfeeding, PLOS ONE 7 (2012) 37219. [2] G. Winger, J.H. Woods, F.G. Hofmann, A Handbook on Drug and Alcohol Abuse, 4th ed., Oxford University Press, Oxford, 2004. [3] E. Merlo-Pich, C. Chiamulera, L. Carboni, Molecular mechanisms of the positive reinforcing effect of nicotine, Behav. Pharmacol. 10 (1999) 587–596. [4] P.R. Kramer, G. Guan, P.J. Wellman, L.L. Bellinger, Nicotine’s attenuation of body weight involves the perifornical hypothalamus, Life Sci. 81 (2007) 500–508. [5] S.S. Audi, M.E. Abraham, A.S. Borker, Effect of cigarette smoke on body weight, food intake and reproductive organs in adult albino rats, Indian J. Exp. Biol. 44 (2006) 562–565.
[6] C. Bishop, G.C. Parker, D.V. Coscina, Systemic nicotine alters wholebody fat utilization in female rats, Physiol. Behav. 80 (2004) 563–567. [7] M. Rezvanipour, S. Esmaeili-Mahani, A. Siahposht, S. Rezvanipour, The effects of morphine and nicotine co-administration on body weight, food intake and appetite-regulating peptides in rats, Iran. J. Vet. Res. (Shiraz Univ.) 12 (2011) 16–23. [8] N. Raoufi, M. Piri, A. Moshfegh, M.C. Shahin, Nicotine improves ethanol-induced impairment of memory: possible involvement of nitric oxide in the dorsal hippocampus of mice, Neuroscience 219 (2012) 82–91. [9] R.G. Cooper, T. Magwere, Nitric oxide-mediated pathogenesis during nicotine and alcohol consumption, Indian J. Physiol. Pharmacol. 52 (2008) 11–18. [10] A. Ledo, J. Frade, R.M. Barbosa, J. Laranjinha, Nitric oxide in brain: diffusion, targets and concentration dynamics in hippocampal subregions, Mol. Asp. Med. 25 (2004) 75–89. [11] S. Pogun, S. Demirgorena, D. Taskirana, L. Kanita, O. Yilmaza, O.E. Koylu, B. Balkan, D. Edythe, Nicotine modulates nitric oxide in rat brain, Eur. Neuropsychol. Pharmacol. 10 (2000) 463–472. [12] C. Han, Q. Zhao, B. Lu, The role of nitric oxide signaling in food intake; insights from the inner mitochondrial membrane peptidase 2 mutant mice, Redox Biol. 1 (2013) 498–507. [13] A.E. Rigamonti, S.G. Cella, G.M. Cavallera, R. Deghenghi, V. Locatelli, N. Pitsikas, E.E. Muller, Contrasting effects of nitric oxide on food intake and GH secretion stimulated by a GH-releasing peptide, Eur. J. Endocrinol. 144 (2001) 155–162. [14] G. Calapai, F. Corica, A. Allegra, A. Corsonello, L. Sautebin, T. De Gregorio, M. Di Rosa, G. Costantino, M. Buemi, A.P. Caputi, Effects
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Effects of chronic nicotine administration on body weight, food intake and nitric oxide concentration in female and male rats Omamuyovwi Meashack Ijomone a,b,∗ , Olayemi Kafilat Olaibi a , Polycarp Umunna Nwoha a b
a Department of Anatomy and Cell Biology, Obafemi Awolowo University, Ile-Ife, Osun, Nigeria Department of Human Anatomy, Cross River University of Technology, Okuku, Cross River, Nigeria
Abstract Nicotine is readily consumed through cigarettes; however it is also easily consumed through the various forms of non-prescription nicotine replacement therapy. It has been shown to possess potential therapeutic value for the management of neurologic and neurodegenerative diseases in the last decade. Hence, this study examined the effects of chronic subcutaneous nicotine administration on food intake and body weight as well as on nitric oxide concentrations and total antioxidant capacity in female and male rats. Nicotine was administered to rats via subcutaneous injections at doses of 0.25, 2 and 4 mg/kg body weight for 28 days. Control groups received normal saline; the vehicle for nicotine. Food intake by each group was monitored daily and body weight of the animals was measured twice weekly. At the end of drug administration, blood was obtained from each animal via cardiac puncture for biochemical determination of serum total antioxidant capacity (TAC) and nitric (NO) concentrations using standard assay kits. Results show significant loss (p < 0.05) of body weight in all nicotine treated female rats. In contrast, male rats showed weight gain, though this was significantly lower (p < 0.001) in nicotine treated groups compared to control. Nicotine significantly reduced (p < 0.001) food consumed in both female and male rats; however dose related changes were observed in only male rats. No significant difference was observed in TAC following nicotine treatments for both female and male rats. Furthermore, only males exhibited changes in NO concentrations following nicotine treatment, as it significantly increased (p < 0.01) NO concentrations in all male treated groups. In conclusion, this study has shown that modulation of body weight, food consumption and nitric oxide formation by nicotine is sexually dimorphic. Also, the study suggests that nicotine modulation of food intake and body weight and its modulation of NO may be independent of each other. © 2014 Elsevier Ireland Ltd. All rights reserved.
Keywords: Nicotine; Body weight; Food intake; Nitric oxide; Sexual differences
1. Introduction Nicotine the natural alkaloid present in Nicotiana tabacum is a readily available substance. It is the main component of cigarettes. However it is also easily consumed through the various forms of non-prescription nicotine replacement therapy such as nicotine dermal patches and nicotine chewing gums [1]. Concerns continually exist on the global effects of nicotine exposure in the body; nevertheless it is well known ∗ Corresponding author at: Department of Anatomy and Cell Biology, Obafemi Awolowo University, Ile-Ife, Osun, Nigeria. Tel.: +234 7031354971. E-mail addresses: [email protected], [email protected] (O.M. Ijomone).
http://dx.doi.org/10.1016/j.pathophys.2014.08.003 0928-4680/© 2014 Elsevier Ireland Ltd. All rights reserved.
that many of the detrimental effects associated with smoking are mostly due to the combination of many other constituents of cigarette smoke, and not nicotine in itself [2]. Though nicotine is the major reason smoking can be addictive, because of its stimulation of the reward-based system within the brain [3], the toxic nature of the many other constituents of cigarettes and their reactions, is largely responsible for serious illness such as coronary heart disease, emphysema and varying forms of cancer caused by smoking [2]. Nevertheless, when administered alone especially by transdermal patches or intravenously, nicotine is known to produce only little or no health problems [2]. Reports indicate an inverse relationship between smoking and body weight, which appears to be mediated by nicotine [4–6]. It has been suggested that nicotine modifies body
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weight via suppression of food intake possible by modulating levels of neuroregulatory substances known to activate or suppress feeding [5,7]. Nicotine has also been seen to have regulatory actions on nitric oxide (NO) synthesis. NO, a neural messenger in the central and peripheral nervous systems is generated from l-arginine by nitric oxide synthase (NOS), following the activation of NMDA receptors [8,9]. In neurons, NO synthesis occurs in an intricate sequence of events that involves stimulation of NMDA-subtype glutamate recetors and influx of Ca2+ into the cytoplasm which upon binding to calmodulin activates neuronal NOS (nNOS) [10]. Stimulation of nAChRs by nicotine is known to activate NMDA receptors and increase intracellular Ca2+ , thus resulting in formation of NO [10,11]. On the other hand, NO has been shown to modulate feeding behaviours. Inhibition of NO synthesis reduces food intake and body weight [12,13]. l-Arginine from which NO is generated results in increased food intake and antagonizes leptin reduction in food intake and body weight [14,15]. Nicotine has been the subject of potential therapeutic value for the management of neurologic and neurodegenerative diseases in the last decade [16,17]. In this study, we have examined the effects of chronic subcutaneous nicotine administration on food intake and body weight as well as on nitric oxide concentrations and total antioxidant capacity. We have used rats of both sexes with a view of appraising sexual dimorphic responses. 2. Materials and methods 2.1. Animal management and treatment Female and male adult albino strain Wistar rats (150–200 g) were used for this study. Animals were housed in clean plastic cages in a clean environment of 12 h day/light cycle, at room temperature. Animals in all groups were allowed access to standard laboratory rat chow and water ad libitum. All experimental protocols were in strict accordance with the guidelines for animal research, as detailed in the NIH Guidelines for the Care and Use of Laboratory Animals (National Academy of Sciences and National Institutes of Health Publications, 2011) and approved by local Institutional Research Committee. A total of 40 rats (20 females and 20 males) were randomly grouped into 4 groups of 5 rats each; a control group and 3 treated groups each for both female and male rats. Treated groups were administered nicotine via subcutaneous injections at doses of 0.25, 2 and 4 mg/kg body weight for 28 days. Control groups received normal saline – vehicle for nicotine. Nicotine was obtained in free base form as (−)nicotine also called (−)-1-methyl-2-(3-pyridyl)pyrrolidine from Sigma Chemicals, USA. Food intake by each group was monitored daily. Weekly or total food consumed was expressed as mean of daily consumptions for each week or throughout the 28-day period of administration. Body weight of the animals was measured twice weekly. Weight gain/loss
was determined by subtracting the body weight at the beginning of administration (initial body weight) from the body weight at end of administration (final body weight). At the end of the administration, animals were euthanized and blood was obtained from each animal via cardiac puncture for biochemical determination of serum TAC and NO concentrations using standard assay kits. 2.2. Biochemical determination of TAC and NO concentrations TAC was determined using the QuantiChromTM Antioxidant Assay Kit (BioAssays Systems, USA). This BioAssay Systems’ improved assay measures total antioxidant capacity in which Cu2+ is reduced by antioxidant to Cu+ . The resulting Cu+ specifically forms a coloured complex with a dye reagent. NO was determined using the QuantiChromTM Nitric Oxide Assay Kit (Bioassays Systems, USA) for the Colorimetric determination of total nitrite and based on the enzymatic conversion of nitrate to nitrite by nitrate reductase. BioAssay Systems’ QuantiChromTM Nitric Oxide Assay Kit is designed to accurately measure NO production following reduction of nitrate to nitrite using improved Griess method. These biochemical studies were carried out following protocols in manufacturer’s guidelines. 2.3. Statistical analysis Data obtained were analyzed using One-way ANOVA, followed by Student Newman–Keuls (SNK) for post-tests. GraphPad Prism 5 (Version 5.03, GraphPad Software, USA.) was the statistical package used for data analysis. Student’s t-test was employed for analysis of between group differences, when appropriate. Statistical significance was set at p < 0.05. 3. Results 3.1. Food intake and body weight The present study observed a marked decrease in weekly body weight in all nicotine treated female groups. Consequently, there was a significant loss of weight in all nicotine treated female groups compared to control (p < 0.05) (Fig. 1). In contrast, male rats showed a steady rise in weekly body weight for all nicotine treated groups, however weight gain was significantly lowered in all nicotine treated male groups compared to control (p < 0.001) (Fig. 1). No dose related changes in body weight gain/loss was observed for both female and male nicotine treated groups. Total food consumed by female rats was significantly reduced in all nicotine treatment groups compared to control (p < 0.001) (Fig. 2). However, no dose related changes were observed. Contrastingly, male rats showed dose related changes following nicotine treatment. Total food consumed by male rats was significantly reduced in all nicotine
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Fig. 1. Line graph of weekly body weight and weight gain/loss of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. *p < 0.05, **p < 0.01, ***p < 0.001. α, β, and δ – significant difference compared to control, 0.25 mg/kg and between 2 mg/kg and 4 mg/kg respectively. One way ANOVA followed by SNK for post-tests.
treatment groups compared to control (p < 0.001). However, 2 mg/kg (72.93 ± 1.95) and 4 mg/kg (77.18 ± 2.28) treatments significantly reduced food intake compared to 0.25 mg/kg (82.93 ± 1.32) nicotine treatment (p < 0.001) (Fig. 2). 3.2. TAC and NO concentrations No significant difference was observed in TAC following nicotine treatments for both male and female rats. Also, no significant difference was observed in NO concentrations of nicotine treated female rats compared to control. In contrast, NO concentrations were significantly increased in all nicotine treatment groups compared to control in male rats (p < 0.01). However no dose related changes were observed (Fig. 3).
4. Discussion The present study showed that chronic subcutaneous administration of nicotine produced sexual dimorphic response in body weight and food consumed. Nicotine resulted in weight loss in females but produced only a slight weight gain in males. Also, nicotine produced dose dependent reduction in total food consumed in males; however, reduction in total food consumed was not dose dependent in females. Reduction in body weight and food intake is a
more consistent effect associated with nicotine administration and tobacco smoking. Numerous studies in both human and animal studies have demonstrated that nicotine and tobacco suppresses body weight and food intake [4–6,18–20]. Chronic nicotine administration has been shown to decrease body weight and food intake [6,7], supporting data from the present study. It has been argued that changes in body weight induced by nicotine results from changes in food intake [6,7], as also supported by this study. In addition, nicotine has been shown to increase lipolysis as well as increase fat utilization. Hence, it has been suggested that nicotine induced changes in whole-body metabolism may also contribute to its weight modulating effects [6]. However, this study has revealed sexual difference in nicotine modulation of body weight and food consumption. Fat deposits are greater in females than males, hence it is possible that nicotine produces a higher fat utilization in females than males and this may contribute to its greater reduction of body weight in females. Also, it is likely that sexual difference exists in nicotine modulation of neuroregulatory substances that suppress and activate feeding [5]. The present study revealed no significant changes in TAC following nicotine treatment compared to control in both females and males. This indicates a lack of effect on antioxidants and possibly in oxidative stress. Evidence has shown that nicotine has both pro-oxidants and antioxidants effects [21]. In vitro studies using Chinese hamsters’ ovary cells
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Fig. 2. Weekly and total food intake of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. *p < 0.05, **p < 0.01, ***p < 0.001. α, β, and δ – significant difference compared to control, 0.25 mg/kg and between 2 mg/kg and 4 mg/kg respectively. One way ANOVA followed by SNK for post-tests.
showed than nicotine induced oxidative stress and resulted in generation of ROS and hydrogen peroxide [22,23]. However, another study demonstrated that nicotine did not show antioxidant properties nor altered indicators of oxidative in vivo and in vitro [24]. Nicotine has also been demonstrated to possess antioxidant properties in experimentally induced neurodegenerative diseases such as Parkinson’s, Alzheimer’s and Huntington’s diseases [25–27]. This has prompted suggestion that its neuroprotective mechanisms may also include antioxidant actions [28]. Dose related effects have also been linked to nicotine dual-actions on oxidative stress. Generally, high doses may stimulate oxidative stress, while low doses may act as antioxidants [29]. The present findings suggest neither antioxidant effect nor induction of oxidative stress. This may not be unconnected to the recent evidence that adult rats are less sensitive to nicotine induced oxidative stress, compared to young or aged ones [30]. The present study also revealed significant increase in serum NO concentrations in all nicotine treated groups compared to control in males only but not in females. Acute or chronically administered nicotine has been demonstrated to increase NO formation in nervous system as well as in systemic organs [9,11]. Studies have reported that NO is important for synaptic plasticity and long-term potentiation (LTP) as well various cognitive and non-cognitive effects [8].
Nicotine is known to improve cognitive processes such as rapid information processing, attention, working memory and long term memory [31,32]. Increasing NO concentrations has been shown as a mechanism by which nicotine improves cognitive functions [8]. However, it is likely that this modulation of NO by nicotine and its resultant benefits may be seen only in males but not females. Also, in the present study, increased NO concentrations in male rats did not affect nicotine reduction of food intake and body weight. This is quite interesting considering that increase in NO synthesis causes increased food intake and body weight [12,14]. Hence, we suggest that nicotine modulation of food intake and body weight may be independent of NO pathways. In summary, there is growing evidence that sexually dimorphic responses are associated with nicotine exposure. These differences are important to consider in developing better treatments for smoking cessation as well as developing forms of nicotine therapy in treatment of neurodegenerative and neurologic disorders. In fact, studies have shown that women find it more difficult to quit smoking than men, and also are less likely to benefit from the use of nicotine replacement therapy in smoking cessation programmes (Perkins et al. [33,34]). Our results support this sexually dimorphic response of nicotine in respect to modulation of body weight, food consumption and nitric oxide formation. One hypothesis
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Fig. 3. TAC and NO concentrations of control and nicotine treated female and male rats. Values are expressed as mean ± SEM; n = 5. **p < 0.01. α – significant difference compared to control. One way ANOVA followed by SNK for post-tests.
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