Chronic Cocaine Administration Increases CNS Tyrosine Hydroxylase Enzyme Activity and mRNA Levels and Tryptophan Hydroxylase Enzyme Activity Levels

Cocaine is an inhibitor of dopamine and serotonin reuptake by synaptic terminals and has potent reinforcing effects that lead to its abuse. Tyrosine hydroxylase (TH) and tryptophan hydroxylase (TPH) catalyze the rate-limiting steps in dopamine and serotonin biosynthesis, respectively, and are the subject of dynamic regulatory mechanisms that could be sensitive to the actions of cocaine. This study assessed the effects of chronic cocaine on brain TH and TPH activities. Cocaine was administered (0.33 mg/infusion, i.v.) to rats for 7 days every 8 min for 6 h per day. This administration schedule is similar to patterns of self-administration by rats when given ad libitum access to this dose. This chronic, response-independent administration increased TH enzyme activity in the substantia nigra (30%) and ventral tegmental area (43%). Moreover, TH mRNA levels were also increased (45 and 50%, respectively). In contrast to the enzymatic and molecular biological changes in the cell bodies, TH activity was unchanged in the terminal fields (corpus striaturn and nucleus accumbens). Similarly, TPH activity was increased by 50% in the raphe nucleus (serotonergic cell bodies). In summary, the chronic response-independent administration of cocaine produces increases in the expression of TH mRNA and activity in both the cell bodies of motor (nigrostriatal) and reinforcement (mesolimbic) dopamine pathways. These increases are not manifested in the terminal fields of these pathways.     

Region - specific alterations in tyrosine hydroxylase activity in rats exposed to lead

Previous studies from our laboratory showed that subchronic exposure to low levels of Pb resulted in significant decrease in dopamine (DA) content, attenuation of stimulus-induced release of DA in the dopaminergic projection area of nucleus accumbens (NA), and alterations in tyrosine hydroxylase (TH) activity in rat whole brain homogenates. The present study reported here was conducted to assess the functional integrity of DA synthesis in different brain regions of rats subchronically (90-days) exposed to 50 ppm Pb by measuring the activity of the rate limiting enzyme, tyrosine hydroxylase, in seven brain regions. In Pb-exposed rats, TH activity was reduced in two of the seven brain regions investigated, i.e., nucleus accumbens (42% reduction) and frontal cortex (61% reduction) when compared to controls. In contrast, Pb exposure did not affect the TH activity in cerebellum, brainstem, hippocampus, hypothalamus and striatum. The changes in TH activity in nucleus accumbens (NA) and frontal cortex (FC) in Pb-exposed rats were further confirmed by Western blot analysis using TH polyclonal antibody. Collectively, these results indicate that low level subchronic Pb exposure may affect TH protein in these brain regions.     

Increases in cytoplasmic dopamine compromise the normal resistance of the nucleus accumbens to methamphetamine neurotoxicity

Methamphetamine (METH) is a neurotoxic drug of abuse that damages the dopamine (DA) neuronal system in a highly delimited manner. The brain structure most affected by METH is the caudate–putamen (CPu) where long-term DA depletion and microglial activation are most evident. Even damage within the CPu is remarkably heterogenous with lateral and ventral aspects showing the greatest deficits. The nucleus accumbens (NAc) is largely spared of the damage that accompanies binge METH intoxication. Increases in cytoplasmic DA produced by reserpine, l -DOPA or clorgyline prior to METH uncover damage in the NAc as evidenced by microglial activation and depletion of DA, tyrosine hydroxylase (TH), and the DA transporter. These effects do not occur in the NAc after treatment with METH alone. In contrast to the CPu where DA, TH, and DA transporter levels remain depleted chronically, DA nerve ending alterations in the NAc show a partial recovery over time. None of the treatments that enhance METH toxicity in the NAc and CPu lead to losses of TH protein or DA cell bodies in the substantia nigra or the ventral tegmentum. These data show that increases in cytoplasmic DA dramatically broaden the neurotoxic profile of METH to include brain structures not normally targeted for damage by METH alone. The resistance of the NAc to METH-induced neurotoxicity and its ability to recover reveal a fundamentally different neuroplasticity by comparison to the CPu. Recruitment of the NAc as a target of METH neurotoxicity by alterations in DA homeostasis is significant in light of the important roles played by this brain structure.     

Decreased Proline Endopeptidase Activity in the Basal Ganglia in Huntington''s Disease

Soluble proline endopeptidase (EC 3.4.21.26) activity was measured by a fluorometric assay in eight human brain areas (caudate nucleus, lateral globus pallidus, medial globus pallidus, substantia nigra-zona compacta, substantia nigra-zona reticulata, frontal cortex-Brodmann area 10, temporal cortex-Brodmann area 38, and hippocampus), in 10 control and 10 Huntington's disease brains. An abnormally low activity (22% of control activity) was found in the caudate nucleus of Huntington's disease brains; significantly decreased activity was also detected in the lateral globus pallidus and medial globus pallidus (37% and 40% of control, respectively).     

Biochemical Evidence for Alteration of Neostriatal Dopaminergic Function by 5,7-Dihydroxytryptamine

Unilateral injection of 5,7-dihydroxytryptamine (DHT) into the rat neostriatum markedly reduced not only striatal tryptophan hydroxylase (TPH) activity but also striatal tyrosine hydroxylase (TH) activity and dopamine (DA) concentration measured 10--15 days later. The decrease in striatal TH activity was dose related over the range of 8--32 micrograms of DHT; a dose of 16 micrograms reduced striatal TH activity to 40--50% of control, DA concentration to 38% of control, and TPH activity to 5--20% of control. Intrastriatal injection of 16 micrograms of DHT reduced TH activity in the ipsilateral substantia nigra to 51% of control. Pretreatment with amfonelic acid, a potent DA uptake inhibitor, significantly reduced the effect of DHT on striatal and nigral TH activity and striatal DA concentration without affecting the DHT-induced decrease in striatal TPH activity. Desmethylimipramine (5 and 25 mg/kg) had no effect on the DHT-induced decrease in striatal TH activity. Striatal choline acetyltransferase and glutamic acid decarboxylase activities were not decreased by 16 micrograms of DHT. The results indicate that DHT can alter dopaminergic function in the rat neostriatum through a direct effect of the drug on DA neurons.     

Melatonin attenuates methamphetamine-induced reduction of tyrosine hydroxylase, synaptophysin and growth-associated protein-43 levels in the neonatal rat brain

Methamphetamine (METH) is a most commonly abused drug which damages nerve terminals by causing formation of reactive oxygen species (ROS), apoptosis, and finally neuronal damage. Fetal exposure to neurotoxic METH causes significant behavioral effects. The developing fetus is substantially deficient in most antioxidative enzymes, and may therefore be at high risk from both endogenous and drug-enhanced oxidative stress. Little is known about the effects of METH on vesicular proteins such as synaptophysin and growth-associated protein 43 (GAP-43) in the immature brain. The present study attempted to investigate the effects of METH-induced neurotoxicity in the dopaminergic system of the neonatal rat brain. Neonatal rats were subcutaneously exposed to 5–10 mg/kg METH daily from postnatal day 4–10 for 7 consecutive days. The results showed that tyrosine hydroxylase enzyme levels were significantly decreased in the dorsal striatum, prefrontal cortex, nucleus accumbens and substantia nigra, synaptophysin levels decreased in the striatum and prefrontal cortex and growth-associated protein-43 (GAP-43) levels significantly decreased in the nucleus accumbens of neonatal rats. Pretreatment with 2 mg/kg melatonin 30 min prior to METH administration prevented METH-induced reduction in tyrosine hydroxylase, synaptophysin and growth-associated protein-43 protein levels in different brain regions. These results suggest that melatonin provides a protective effect against METH-induced nerve terminal degeneration in the immature rat brain probably via its antioxidant properties.     

Morphine and Cocaine Exert Common Chronic Actions on Tyrosine Hydroxylase in Dopaminergic Brain Reward Regions

We studied levels of tyrosine hydroxylase immunoreactivity and phosphorylation state in the ventral tegmental area (VTA) and nucleus accumbens (NAc) in an effort to understand better the mechanisms by which these brain reward regions are influenced by opiates and cocaine. In the VTA, chronic, but not acute, administration of either morphine or cocaine increased levels of tyrosine hydroxylase immunoreactivity by 30-40%, with no change observed in the relative phosphorylation state of the enzyme. In the NAc, chronic, but not acute, morphine and cocaine treatments decreased the phosphorylation state of tyrosine hydroxylase, without a change in its total amount. In contrast, morphine and cocaine did not regulate tyrosine hydroxylase in the substantia nigra or caudate/putamen, brain regions generally not implicated in drug reward. Morphine and cocaine regulation of tyrosine hydroxylase could represent part of a common biochemical basis of morphine and cocaine addiction and craving.     

氨甲酰胆碱引起的促钠排泄与下丘脑TH—IR神经元活性的变化

目的：我们最近的实验发现大鼠侧脑室注射氨甲酰胆碱引起显著的促钠排泄作用,本工作同时还观察了下丘脑内不同脑区的儿茶酚胺能神经元活性的变化。方法和结果：氨甲酰胆碱注射后４０ｍｉｎ,下丘脑室旁核的腹侧和内侧小细胞部、内侧视前区、尾核、苍白球的酪氨酸羟化酶免疫反应（ｔｈｙｒｏｓｉｎｅｈｙｄｒｏｘｙｌａｓｅｉｍｍｕｎｏｒｅａｃｔｉｖｉｔｙ,ＴＨＩＲ）阳性细胞数减少,免疫反应染色强度降低;下丘脑室旁核的后部,下丘脑前区的后部、下丘脑室周核、弓状核、下丘脑外侧区的ＴＨＩＲ阳性细胞数增多,免疫反应染色强度增强。结论：侧脑室注射氨甲酰胆碱对脑内不同脑区的内源性儿茶酚胺能神经元分别有兴奋或抑制作用,其与促钠排泄的关系将在本文中讨论     

The relationship between the proteolytic activity in the basal nuclei and locomotion in the open-field test under the conditions of an altered photoperiod

The effect of an altered photoperiod on the proteolytic activity in the basal nuclei, including the caudate nucleus, globus pallidus, nucleus accumbens, and amygdala complex, and rat behavior in the openfield test was studied. The altered photoperiod modulated the functional state of the basal nuclei, which was reflected in the locomotor activity of the animals.

     

Effect of stimulation of the caudate nucleus on activity of neurons of the globus pallidus

Activity of neurons of the globus pallidus was recorded extracellularly during stimulation of the caudate nucleus. It is demonstrated that background activity (BA) of most neurons of the globus pallidus is depressed under these conditions, which is regarded as a manifestation of inhibition of the investigated neurons. The period of BA depression varied in different cells from 60 to 500 msec. In some cases this period was preceded by emergence of an action potential with a latent period of 10–20 msec. In addition to inhibition of the activity of globus pallidus neurons during stimulation of the caudate nucleus, it was possible to record evoked responses of the given neurons in the form of group discharges with a latent period of 18–40 msec and single action potentials with a latent period of 50–100 msec. The neurons that reacted with a shorter latent period were localized at the lateral limit of the globus pallidus, whereas neurons with other kinds of responses were distributed in the globus pallidus comparatively evenly.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 1, No. 2, pp. 202–209, September–October, 1969.     

Effect of TRH analog (DN-1417) on tyrosine hydroxylase activity in mesocortical, mesolimbic and nigrostriatal dopaminergic neurons of rat brain

Tyrosine hydroxylase (TH) was assayed in eight regions of rat brain following repeated treatment with a TRH analog, DN-1417 (gamma-butyrolactone-gamma-carbonyl-histidyl-prolinamide). Repeated DN-1417 treatment (20 mg/kg/day, IP) for 7 days increased TH activity in the ventral tegmental area and decreased in the prefrontal cortex polar, medial and lateral fields and olfactory tubercles. No significant change in TH activity was found in the nucleus accumbens, striatum and substantia nigra. Kinetic analysis showed that the increased TH activity in the ventral tegmental area was due to an increase in Vmax, but not a change in the apparent Km of TH for a cofactor, 6-methyl-tetrahydropteridine. When TH was assayed at a suboptimal pH and in the presence of a subsaturating cofactor, the striatal TH was activated significantly after DN-1417. In the prefrontal cortex medial field, nucleus accumbens and olfactory tubercles, TH activity assayed under the suboptimal condition was not modified by DN-1417 treatment. These results suggest an intimate involvement of central dopaminergic systems in the actions of DN-1417.     

Differential Changes in the Content of Amino Acid Neurotransmitters in Discrete Regions of the Rat Brain Prior to the Onset and During the Course of Homocysteine-Induced Seizures

Changes in amino acid concentrations were investigated in selected regions of rat brain prior to the onset and during the course of epileptiform seizures induced by L-homocysteine. The concentration of gamma-aminobutyric acid (GABA) decreased preictally in substantia nigra (-18%), caudate putamen (-26%), and inferior colliculus (-46%). After seizure onset, the GABA content was further reduced in substantia nigra (-31%) and additionally in hippocampus (-18%). Preictal taurine levels were elevated in globus pallidus (+26%) and caudate putamen (+13%) but returned to normal after seizure onset. However, in hippocampus, taurine decreased both preictally (-22%) and after seizure onset (-56%). Glycine was reduced preictally only in globus pallidus (-13%). After seizure onset the direction of its concentration change varied in the brain regions studied. Glutamate levels decreased preictally in hippocampus (-10%) and hypothalamus (-46%) but increased in globus pallidus (+14%). Normal levels were detectable after seizure onset in hypothalamus and globus pallidus but a further reduction in hippocampus (-59%) and significant reductions in substantia nigra (-15%) and caudate putamen (-17%) were detected. Aspartate was elevated in hippocampus, both preictally (+49%) and after seizure onset (+21%) while at the same phases in globus pallidus a consistent reduction (-30%) was observed. The glutamine content increased preictally in globus pallidus (+41%) and hypothalamus (+36%), and in all brain areas during the ictal phase of seizure, the hippocampus exhibiting a dramatic increase (approximately 300%). The contents of serine and alanine were altered in most regions studied only after seizure onset, with the exception of the hippocampus, where a decrease (-41%) of serine was observed preictally.     

Huntington's disease: regional alteration in muscarinic cholinergic receptor binding in human brain.

Huntington's Disease, an autosomal dominant neurological disorder, is characterized by diffuse neuronal degeneration particularly in the basal ganglia and cerebral cortex. The purpose of this study was to examine various discrete regions of choreic and control brains for alterations in muscarinic cholinergic receptor binding and choline acetyltransferase (ChAc) activity. Nine postmortem brains, three from patients with Huntington's Disease and six controls, were dissected into 17 discrete regions. Each regional homogenate was assayed for muscarinic receptor concentration by measuring specific membrane binding of [³H]-QNB, a potent muscarinic antagonist which selectively labels brain muscarinic receptors. Aliquots from each brain region were also assayed for ChAc activity. Of significance was the marked reduction in specific [³H]-QNB receptor binding in the caudate nucleus, putamen and globus pallidus of choreic brain while no significant alterations were detected in other brain regions. Significant decreases in ChAc activity were found in the caudate nucleus, putamen, and globus pallidus with no alterations in ChAc activity in the rest of the brain regions examined. The tissues were chosen such that protein levels were similar in both choreic and normal brain samples. The apparent reduction in the number of muscarinic cholinergic receptors in the choreic brains suggests that treatment with cholinomimetic drugs might be beneficial in Huntington's Disease.     

Acute administration of cocaine regulates the phosphorylation of serine-19, -31 and -40 in tyrosine hydroxylase

Acute cocaine can inhibit catecholamine biosynthesis by regulating the enzymatic activity of tyrosine hydroxylase via alterations in the phosphorylation state of the enzyme. The mechanisms underlying acute cocaine-dependent regulation of tyrosine hydroxylase phosphorylation have not been determined. In this study, 0, 15 or 30 mg/kg cocaine was administered intraperitoneally to rats and the phosphorylation state of tyrosine hydroxylase in the brain was examined using antibodies specific for the phosphorylated forms of serine-19, -31 and -40 in tyrosine hydroxylase. In the caudate and nucleus accumbens, cocaine dose-dependently decreased the levels of phosphorylated serine-19, -31 and -40. In the ventral tegmental area, the levels of phosphorylated serine-19, but not serine-31 and -40, were decreased by 15 and 30 mg/kg cocaine. In the amygdala, the levels of phosphorylated serine-19, but not serine-31 or -40, were decreased. The functional effects of these alterations in phosphorylation state were assessed by measuring tyrosine hydroxylase activity in vivo (accumulation of DOPA after administration of the decarboxylase inhibitor NSD-1015). Acute administration of 30 mg/kg cocaine significantly decreased l-DOPA production in caudate and accumbens but not in amygdala. These data suggest that the phosphorylation of serine-31 or -40, but not serine-19, is involved in the regulation of tyrosine hydroxylase activity by acute cocaine.     

The Dopamine D1-D2 Receptor Heteromer Localizes in Dynorphin/Enkephalin Neurons: INCREASED HIGH AFFINITY STATE FOLLOWING AMPHETAMINE AND IN SCHIZOPHRENIA*

The distribution and function of neurons coexpressing the dopamine D1 and D2 receptors in the basal ganglia and mesolimbic system are unknown. We found a subset of medium spiny neurons coexpressing D1 and D2 receptors in varying densities throughout the basal ganglia, with the highest incidence in nucleus accumbens and globus pallidus and the lowest incidence in caudate putamen. These receptors formed D1-D2 receptor heteromers that were localized to cell bodies and presynaptic terminals. In rats, selective activation of D1-D2 heteromers increased grooming behavior and attenuated AMPA receptor GluR1 phosphorylation by calcium/calmodulin kinase IIα in nucleus accumbens, implying a role in reward pathways. D1-D2 heteromer sensitivity and functional activity was up-regulated in rat striatum by chronic amphetamine treatment and in globus pallidus from schizophrenia patients, indicating that the dopamine D1-D2 heteromer may contribute to psychopathologies of drug abuse, schizophrenia, or other disorders involving elevated dopamine transmission.     

Neurotransmitter receptor alterations in Parkinson's disease.

Neurotransmitter receptor binding for GABA, serotonin, cholinergic muscarinic and dopamine receptors and choline acetyltransferase (ChAc) activity were measured in the frontal cortex, caudate nucleus, putamen and globus pallidus from postmortem brains of 10 Parkinsonian patients and 10 controls. No changes in any of these systems were observed in the frontal cortex. In the caudaye nucleus, only the apparent dopamine receptor binding was altered with a significant 30% decrease in the Parkinsonian brain. Both cholinergic muscarinic and serotonin receptor binding were significantly altered in the putamen, the former increasing and the latter decreasing with respect to controls. In addition, ChAc activity was decreased in the putamen. In the globus pallidus, only ChAc activity was significantly changed, decreasing about 60%, with no change in neurotransmitter receptor binding. The results suggest that a progressive loss of dopaminergic receptors in the caudate nucleus may contribute to the decreased response of Parkinsonian patients to L-dopa and dopamine agonist therapy.     

Progesterone receptor isoforms differentially regulate the expression of tryptophan and tyrosine hydroxylase and glutamic acid decarboxylase in the rat hypothalamus

Progesterone exerts a variety of actions in the brain through the interaction with its receptors (PR) which have two isoforms with different function and regulation: PR-A and PR-B. Progesterone may modulate neurotransmission by regulating the expression of neurotransmitters synthesizing enzymes or their receptors in several brain regions. The role of PR isoforms in this modulation is unknown. We explored the role of PR isoforms in the regulation of tryptophan (TPH) and tyrosine (TH) hydroxylase, and glutamic acid decarboxylase (GAD) expression in the hypothalamus of ovariectomized rats. Two weeks after ovariectomy, animals were subcutaneously injected with 5 μg of estradiol benzoate (EB), and 40 h later, progesterone (P) was intracerebroventricularly (ICV) injected. Each animal received two ICV injections of 1 μg/μl (4 nmol) of PR-B and total PR (PR-A + PR-B) sense or antisense (As) oligonucleotides (ODNs). First injection was made immediately before sc EB injection, and 24 h later animals received the second one. Twenty-four hours after P administration, rats were euthanized and brains removed to measure the expression of PR-A and PR-B, TPH, TH and GAD by Western blot. We observed that sense ODNs modified neither PR isoforms nor enzymes expression in the hypothalamus, whereas PR A + B antisense (PR A + B As) clearly decreased the expression of both PR isoforms in this region. ICV administration of PR-B As only decreased PR-B isoform expression with no significant effects on PR-A expression. A differential protein expression of TPH, TH and GAD was observed after PR isoforms antisense administration. PR-B As administration decreased the expression of TPH (65% with respect to control). In contrast, PR A + B As and PR-B As administration increased (51.6% and 34.4%, respectively) TH expression. The administration of PR A + B As and PR-B As diminished GAD expression (33.4% and 41.6%, respectively). Our findings indicate that PR isoforms play a differential role in the regulation of the content of TPH, TH and GAD in the rat hypothalamus.     

Role of the caudate nucleus in the development of evoked synchronized activity

Synchronized activity (spindles, augmentation response) evoked by stimulation of thalamic nonspecific, association, and specific nuclei was investigated in chronic experiments on 11 cats before and after successive destruction of the caudate nuclei. After destruction of the caudate nuclei the duration of spindle activity in the frontal cortex and subcortical formations (thalamic nuclei, globus pallidus, putamen) was reduced to only three or four oscillations. In the subcortical nuclei its amplitude fell significantly (by 50±10%); in the cortex the decrease in amplitude was smaller and in some cases was not significant. Different changes were observed in the amplitude of the augmentation response, depending on where it was recorded. In the subcortical formations it was considerably and persistently reduced (by 50±10%); in the cortex these changes were unstable in character. Unilateral destruction of the caudate nucleus inhibited synchronized activity evoked by stimulation of the thalamic nuclei on the side of the operation only. Destruction of the basal ganglia (caudate nucleus, globus pallidus, entopeduncular nucleus, and putamen) did not prevent the appearance of synchronized activity; just as after isolated destruction of the caudate nucleus, after this operation synchronized activity was simply reduced in duration and amplitude. It is suggested that the caudate nucleus exerts an ipsilateral facilitatory influence on the nonspecific system of the thalamus during the development of evoked synchronized activity.A. A. Bogomolets Institute of Physiology, Academy of Sciences of the Ukrainian SSR, Kiev. Translated from Neirofiziologiya, Vol. 9, No. 3, pp. 239–248, May–June, 1977.     

Tyrosine Hydroxylase Activity in Caudate Nucleus from Parkinson''s Disease: Effects of Iron and Phosphorylating Agents

Tyrosine hydroxylase (TH) activity of human postmortem brain tissues from controls and patients with Parkinson's disease (PD) was examined in the presence of Fe2+ and phosphorylation agents, such as cyclic AMP, exogenous protein kinase, calcium plus calmodulin (Ca2+-CaM), and ATP. TH activity from parkinsonian tissue was increased by 48% with statistical significance in the presence of exogenous protein kinase. Cyclic AMP alone had no effect, whereas Ca2+-CaM increased the activity by only 10%. The presence of acetylcholine resulted in a slight decrease in enzyme activity. Human TH was stimulated 13.17-fold in the presence of 1 mM Fe2+. For iron dependence, no significant differences could be shown for the Km values of TH in striata of PD, while the activity of TH was half of that of controls. Here stimulation with 1 mM Fe2+ raised the activity of TH 11-fold. Stimulation of rat, gerbil, pig, and human caudate nucleus TH with Fe2+ shows remarkable species differences. In particular, the sensitivity of human TH to stimulating processes is noteworthy. H2O2 decreases TH activity only at high concentrations. Species differences are noted for the combined incubation of Fe2+ and H2O2. In the gerbil caudate nucleus, H2O2 does not prevent the stimulating properties of Fe2+, while the pig shows a dose-dependent decline of TH activity. In conclusion, there are no significant changes in the stimulating properties of human caudate nucleus TH activity with Fe2+ in PD, while such differences are noted by using exogenous protein kinase. Furthermore, experimental evidence shows that TH activity declines at high concentrations of H2O2 only. Potentiation of this effect by Fe2+ seems to be species-dependent.     

In vivo nicotine treatment regulates mesocorticolimbic CREB and ERK signaling in C57Bl/6J mice

The extracellular regulated kinase (ERK) pathway was studied to determine its role in neuronal plasticity related to the development of nicotine dependence. Levels and phosphorylation state of ERK, cAMP response element binding protein (CREB) and proline-rich/Ca2+-activated tyrosine kinase (PYK2), and levels of tyrosine hydroxylase (TH), were determined using western blotting. C57Bl/6J mice received acute or chronic nicotine (200 microg/mL) in their drinking water or were withdrawn from nicotine for 24 h following chronic exposure. CREB phosphorylation was reduced in the nucleus accumbens following chronic nicotine, consistent with previous reports that decreased accumbens CREB activity increases drug reinforcement. In contrast, CREB phosphorylation was increased in the prefrontal cortex following chronic nicotine exposure and in the ventral tegmental area during nicotine withdrawal. In addition, total and phosphorylated ERK decreased in the amygdala following chronic nicotine exposure, but ERK phosphorylation increased in the prefrontal cortex. TH levels increased in both the amygdala and prefrontal cortex, supporting the hypothesis that increased catecholaminergic tone contributes to nicotine reinforcement. Overall, these results support a role for ERK and CREB activity in neural plasticity associated with nicotine dependence.