Chronic nicotine treatment decreases neurofilament immunoreactivity in the rat ventral tegmental area
Introduction
Chronic addictive drug exposure induces adaptive changes in neural circuits that ultimately modify the function of emotional and motivational processes involved in addictive behavior. In recent years, several authors have shown that particular biochemical substrates (transcription factors, immediate early genes, receptors, G proteins, and kinases) are altered by chronic treatment with either cocaine or morphine at doses that are known to produce changes in rat behavioral outcome, such as locomotor activation, preference for drug solutions, and drug self-administration (Nestler et al., 1993). The neuroanatomical sites specifically affected by chronic cocaine or morphine treatment include the ventral tegmental area and the nucleus accumbens (Nestler et al., 1993), the origin and the main terminal field of the mesolimbic dopamine system, respectively (LeMoal and Simon, 1991). A vast array of data has implicated this system in the reinforcing effects of drugs of abuse Koob, 1992, Kreek and Koob, 1998.
Among the various biochemical changes, reduction in the expression of three of the neural intermediate filament proteins of the type IV family, NF-68, NF-160 and NF-200 (Zuoshang et al., 1994), was described for the ventral tegmental area of rats exposed to chronic treatment with morphine, alcohol or cocaine Beitner-Johnson et al., 1992, Nestler et al., 1994. Parallel biochemical and behavioral studies on biochemical changes in the ventral tegmental area of inbred Lewis and Fischer rat strains have shown that, phenotypically, the former strain resembles the latter on exposure to chronic treatment with morphine, cocaine or alcohol (Nestler et al., 1993). Moreover, naive Lewis rats showed reduced neurofilament protein levels in the ventral tegmental area similar to those described after chronic cocaine or morphine treatment Guitart et al., 1992, Guitart et al., 1993b, Guitart et al., 1993a, supporting the hypothesis that changes in neurofilament proteins are involved in the genetically determined ‘drug-preferred’ state.
These findings suggest that chronic use of drugs of abuse generates long-term changes in the expression of proteins involved in the maintenance of the neuronal cytoskeleton. These changes eventually participate in the neuroadaptation of the activity of the mesolimbic dopaminergic pathway that is believed to act as substrate for addictive behavior Guitart et al., 1993a, Miserendino et al., 1993.
Nicotine, similar to other drugs of abuse, activates the mesolimbic pathway by increasing cell firing of dopaminergic neurons in the ventral tegmental area through activation of nicotinic receptors Calabresi et al., 1989, McGehee et al., 1995. Interestingly, chronic nicotine treatment results in sensitization of dopamine release in response to nicotine microinfusions into the ventral tegmental area (Balfour et al., 1998) in a manner that is reminiscent of the effects of psychostimulants.
The aim of the present study was to investigate the changes of neurofilament proteins in the ventral tegmental area following chronic nicotine treatment according to regimens similar to those known to produce sensitization of dopamine release to challenging doses of nicotine (Balfour et al., 1998). Validated computer-assisted morphologic methods were applied to measure changes in immunoreactivity for NF-68, NF-160, and NF-200 neurofilament proteins in regions of the ventral tegmental area and substantia nigra that contain dopaminergic neurons.
Section snippets
Subjects and drug treatments
Male Wistar rats (initial body weight 170–240 g; Charles River, Calco, Italy) were used. Animals were housed in three or four per cage and maintained in a temperature-controlled environment on a 12-h light–12-h dark cycle with light on at 6:00 a.m. with food pellets and water ad libitum. All animal procedures and research complied with national legislation and with the company policy on the Care of Use of Animals and with related codes of practice.
Chronic nicotine treatment consisted of daily
Results
Visual inspection suggested that immunoreactivity for all neurofilament proteins was reduced in the ventral tegmental area of rats exposed to chronic nicotine when compared to that of rats exposed to saline (Fig. 1a–f). Statistical analysis of the microdensitometric measurements of ventral tegmental areas indicated that the reduction in immunoreactivity was significant for NF-160 (P<0.01) and NF-200 (P<0.05), showing a relative reduction of 44.5% and 22.5%, respectively, when compared to that
Discussion
The data presented in this article demonstrate that exposure to chronic nicotine treatment reduced neurofilament protein immunoreactivity in the ventral tegmental area of Wistar rats. The reduction did not depend on loss of neurons, since the cell count showed no difference between experimental groups. The observed reduction of NF-160 and NF-200 immunoreactivity and the trend to a reduction observed for NF-68 immunoreactivity could be interpreted as the result of negative transcriptional
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