Effects of chronic nicotine administration and its withdrawal on striatal FosB/ΔFosB and c-Fos expression in rats and mice
Introduction
Fos family proteins are transcription factors, and they are believed to act as initiators of long-term cellular changes in response to different kinds of stimulation. They heterodimerize with Jun family transcription factors to form the activator protein-1 (AP-1) complex which can either induce or repress expression of genes (Morgan and Curran, 1991, Chao and Nestler, 2004), and by this way they can induce long-term adaptive changes. c-fos is a prototypical immediate early gene (IEG) and its induction has been used as a marker for postsynaptic activation (Sagar et al., 1988, Morgan and Curran, 1991). Acute administration of nicotine induces the expression of c-Fos rapidly and transiently in various brain areas of rats (Ren and Sagar, 1992, Kiba and Jayaraman, 1994, Salminen et al., 1996, Salminen et al., 1999, Seppä et al., 2001). However, during continuous administration of nicotine we found that the induction of c-Fos was attenuated (Salminen et al., 1999). On the other hand, repeated nicotine injections inducing behavioural sensitization in rats enhanced the accumbal expression of c-Fos (Mathieu-Kia et al., 1998, Shim et al., 2001).
During chronic administration of abused drugs among which cocaine is the most frequently studied one, another Fos-family protein, ΔFosB, has been shown to accumulate within the brain (Hope et al., 1994, Nye and Nestler, 1996, Pich et al., 1997). ΔFosB is a truncated splice variant of full-length FosB, and lacks a portion of the C-terminal domain present in other Fos proteins (Yen et al., 1991). Because of its long half-life it persists in brain for a longer time than other Fos family transcription factors, and it has been implicated in drug addiction (Chen et al., 1997, Nestler, 2004). Previous studies suggested that ΔFosB might play a critical role in cocaine-induced reward and locomotor activation (Kelz et al., 1999, Colby et al., 2003). Increased ΔFosB expression was found in rats which had self-administered nicotine for 2 weeks (Pich et al., 1997) but the effects of nicotine on the expression of ΔFosB have not been otherwise studied. The main purpose of this study was to investigate the effects of long-term oral nicotine treatment on striatal ΔFosB expression in mice. We have earlier shown that nicotine can be administered chronically in the drinking water to mice in a way that mimics the plasma nicotine pharmacokinetics of human smokers (Pekonen et al., 1993). On the 50th day of this long-term treatment nicotine activates the motor activity of the mice and stimulates their accumbal and striatal dopamine release and metabolism (Pietilä et al., 1995, Gäddnäs et al., 2000, Gäddnäs et al., 2001, Gäddnäs et al., 2002). Therefore, we studied the expression of c-Fos to see whether the “acute” IEGs are induced during this long-term nicotine treatment.
The importance of the mesolimbic dopaminergic system in reinforcement-based behaviours and addiction is well-documented (Wise, 2002), and the nigrostriatal system is also involved in reward-based sensory-motor conditioning and motor responsiveness to addictive drugs (Graybiel, 1995). Furthermore, several drugs of abuse have been shown to induce ΔFosB robustly in the striatum including nucleus accumbens (NAcc) (Nestler, 2004). For these reasons the expression of Fos family proteins was studied in the two major dopaminergic brain areas, NAcc and caudate-putamen (CPu). As a specific antibody for ΔFosB is not available, the ΔFosB expression was studied by using an antibody which does not differentiate between the fosB encoded proteins FosB and ΔFosB. Thus, FosB/ΔFosB expression was studied also after withdrawal to see whether its long-lasting increased expression would point at long-lived ΔFosB. We found that in nicotine-drinking mice the expression of FosB/ΔFosB in the NAcc was elevated on the 50th day of nicotine administration but returned to control levels after 24-h withdrawal. Therefore, we decided to give nicotine repeatedly to rats to see if simultaneously with development of locomotor sensitization long-lasting elevation of ΔFosB expression occurs. Part of this work was presented earlier in abstract form (Marttila et al., 2003a, Marttila et al., 2003b).
Section snippets
Experimental procedures
All animal experiments were conducted according to the “European Convention for the Protection of Vertebrate Animals used for Experimental and other Scientific Purposes”. The experimental design was approved by the Committee for Animal Experiments of the University of Helsinki.
Administration of nicotine to mice
Five-week-old male NMRI mice (body weight 20–25 g in the beginning of experiments) bred locally in the Laboratory Animal Center, University of Helsinki, were divided randomly into nicotine-receiving and control mice.
FosB/ΔFosB and c-Fos IS in nicotine-treated mice
The FosB/ΔFosB expression in the NAcc and CPu of mice on the 50th day with nicotine in the drinking water is shown in Fig. 2. The number of FosB/ΔFosB positive nuclei was significantly elevated (P = 0.013, Student's t-test) in the NAcc of nicotine treated-mice (282 ± 24) as compared with control mice (192 ± 21). There was no significant effect of nicotine on the number of FosB/ΔFosB positive nuclei in the CPu (nicotine-treated mice 124 ± 12; control mice 112 ± 9). To study the duration of the elevated
Discussion
Previous studies have suggested that transcriptional regulation of the fosB gene products plays a critical role in the behavioural responses to addictive drugs and thus, in the development of addiction (for references see Introduction). In the present study FosB/ΔFosB and c-Fos expressions were studied in two dopaminergic brain areas, nucleus accumbens (NAcc) and caudate-putamen (CPu), to clarify the effects of long-term nicotine administration, and consequently the molecular mechanisms of
Acknowledgements
The authors wish to thank Kati Rautio for excellent technical assistance. This study was supported by the Finnish Foundation for Alcohol Studies and by a grant from the Helsinki University Pharmacy to K.M.
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