Jesus A. Angulo

Molecular aspects of neuropeptide regulation and function in the corpus striatum and nucleus accumbens.

In the corpus striatum and nucleus accumbens, neuropeptides participate along with conventional neurotransmitters such as dopamine, gamma-aminobutyric acid (GABA), acetylcholine and glutamate in the regulation of locomotor activity, stereotyped motor behaviors and neural events related to reward and affective state. The present review concerns itself with four major neuropeptide systems--enkephalin, dynorphin, tachykinins and neurotensin--and it summarizes neuroanatomical and functional studies as well as emphasizing regulatory interactions between neurotransmitters and neuropeptides at the level of neuropeptide gene expression. Dopaminergic transmission emanating from midbrain dopaminergic cell bodies of the substantia nigra and the ventral tegmentum regulates striatal and accumbens neuropeptide levels and their mRNAs. Evidence is presented for D1 or D2 receptor involvement as well as D1-D2 interactions that modulate neuropeptide and mRNA levels in striatum and accumbens neurons. Regulatory influences by GABAergic, serotonergic and cortical (glutamatergic) neurotransmission and via sigma receptors and circulating adrenal steroids are also described. The evidence gathered in many laboratories thus far indicates that these major basal ganglia peptidergic systems are modulated dynamically and sometimes in opposing ways by various neurochemical inputs which alter neuropeptide and neuropeptide mRNA levels over both short- and long-term. Neuropeptide systems are involved in the regulation and execution of motor programs and may also be involved in the control of mood and affect as well as self-administration behavior and behavioral sensitization, especially via the nucleus accumbens and its reciprocal connections with the midbrain, hippocampus and frontal cortex. Glucocorticoids modulate mood as well as self-administration behavior and influence locomotor activity and certain forms of stereotypy. The modulation of striatal proenkephalin and protachykinin mRNA levels by adrenal steroids is described along with distribution of adrenal steroid receptor subtypes. Adrenal steroid regulation of neuropeptide gene expression in striatum, accumbens and midbrain suggests that there may be a wider role for glucocorticoids and for other neuropeptide systems in environmental and drug influences on normal and abnormal behaviors involving the nigrostriatal and mesolimic systems.

Comparison of Cocaine‐ and Methamphetamine‐Evoked Dopamine and Glutamate Overflow in Somatodendritic and Terminal Field Regions of the Rat Brain during Acute, Chronic, and Early Withdrawal Conditions

Abstract: Methamphetamine and cocaine are among the most commonly abused psychostimulants. Repeated injections of psychostimulants produce behavioral sensitization or augmented locomotion in rats. Behavioral sensitization to methamphetamine and cocaine is long lasting and persists after cessation of drug treatment. Because dopamine and glutamate are major neurotransmitters of the neostriatum, we evaluated the profile of cocaine‐ or methamphetamine‐evoked dopamine and glutamate overflow in the caudate putamen, nucleus accumbens, ventral tegmental area, and substantia nigra compacta of the rat brain. We also compared acute exposure to these drugs with chronic treatment and early withdrawal. Acute injection of methamphetamine (1 mg/kg of body weight) or cocaine (10 mg/kg) resulted in elevated levels of extracellular dopamine in all brain regions measured, although the magnitude of increase varied between brain regions. Overall, methamphetamine caused more dopamine to accumulate in the extracellular space than did cocaine when administered to animals during early withdrawal (7 days of daily injections and challenge on day 11). For example, a challenge injection of methamphetamine produced a greater elevation of extracellular dopamine in the caudate putamen when compared to acute (naïve) exposure. By contrast, a challenge injection of cocaine resulted in dopamine levels in the caudate putamen that were lower than those observed for acute exposure. In the ventral tegmental area and the substantia nigra compacta, a challenge injection of methamphetamine or cocaine resulted in extracellular dopamine levels that were lower than those for acute exposure. Thus, it appears that behavioral sensitization to cocaine can be sustained during early withdrawal in the absence of augmented drug‐evoked dopamine overflow. Acute injection of methamphetamine or cocaine did not change extracellular levels of glutamate in the neostriatum. Cocaine challenge (early withdrawal) increased glutamate overflow in the caudate putamen and the nucleus accumbens. In contrast, methamphetamine challenge increased glutamate overflow in the caudate putamen, but it decreased glutamate in the nucleus accumbens. In the ventral tegmental area and the substantia nigra compacta, acute methamphetamine exposure decreased glutamate oveflow, but acute cocaine exposure increased it. Although amphetamines and cocaine induce similar behavioral responses, the results presented here demonstrate that at the neurochemical level (neurotransmitter release) they sometimes evoke opposite effects depending on the brain region studied and the duration of drug treatment. Moreover, the sensitized augmentation of locomotor activity observed by us and others in response to a challenge injection of cocaine is not dependent on elevation of the extracellular concentration of dopamine in the neostriatum. We are currently investigating the hypothesis that cocaine activates peptidergic systems of the neostriatum and that these systems modulate the synaptic release of dopamine in response to psychostimulants.

METHAMPHETAMINE-INDUCED CELL DEATH: SELECTIVE VULNERABILITY IN NEURONAL SUBPOPULATIONS OF THE STRIATUM IN MICE

Methamphetamine (METH) is an illicit and potent psychostimulant, which acts as an indirect dopamine agonist. In the striatum, METH has been shown to cause long lasting neurotoxic damage to dopaminergic nerve terminals and recently, the degeneration and death of striatal cells. The present study was undertaken to identify the type of striatal neurons that undergo apoptosis after METH. Male mice received a single high dose of METH (30 mg/kg, i.p.) and were killed 24 h later. To demonstrate that METH induces apoptosis in neurons, we combined terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining with immunohistofluorescence for the neuronal marker neuron-specific nuclear protein (NeuN). Staining for TUNEL and NeuN was colocalized throughout the striatum. METH induces apoptosis in approximately 25% of striatal neurons. Cell counts of TUNEL-positive neurons in the dorsomedial, ventromedial, dorsolateral and ventrolateral quadrants of the striatum did not reveal anatomical preference. The type of striatal neuron undergoing cell death was determined by combining TUNEL with immunohistofluorescence for selective markers of striatal neurons: dopamine- and cAMP-regulated phosphoprotein, of apparent Mr 32,000, parvalbumin, choline acetyltransferase and somatostatin (SST). METH induces apoptosis in approximately 21% of dopamine- and cAMP-regulated phosphoprotein, of apparent Mr 32,000-positive neurons (projection neurons), 45% of GABA-parvalbumin-positive neurons in the dorsal striatum, and 29% of cholinergic neurons in the dorsal-medial striatum. In contrast, the SST-positive interneurons were refractory to METH-induced apoptosis. Finally, the amount of cell loss determined with Nissl staining correlated with the amount of TUNEL staining in the striatum of METH-treated animals. In conclusion, some of the striatal projection neurons and the GABA-parvalbumin and cholinergic interneurons were removed by apoptosis in the aftermath of METH. This imbalance in the populations of striatal neurons may lead to functional abnormalities in the output and processing of neural information in this part of the brain.

Regulation by dopaminergic neurotransmission of dopamine D2 mRNA and receptor levels in the striatum and nucleus accumbens of the rat.

The effect of dopamine depletion or pharmacological blockade of dopamine receptors on striatal and accumbens dopamine D2 mRNA and receptor levels was assessed by in situ hybridization histochemistry and receptor autoradiography. The time course of pharmacological blockade with haloperidol demonstrates a complex mode of regulation of dopamine D2 mRNA and receptor levels. By day 8 of haloperidol treatment, D2 mRNA and receptor levels were decreased (up to 20%) in the medial and anterior aspects of the caudate-putamen (mCPU and aCPU) and the nucleus accumbens (NAc). However, by day 21 of haloperidol treatment, D2 mRNA and receptor were increased relative to vehicle-injected controls. Likewise, unilateral dopamine depletion due to 6-hydroxydopamine (6-OHDA) lesions of mesencephalic dopaminergic neurons resulted in decreased levels of D2 receptor mRNA by day 8 post-lesion in the ipsilateral mCPU, aCPU and the NAc. However, at days 14 or 21 post-lesion, there was a reversal of the effect with increases of up to 22% in all brain regions ipsilateral to the lesion. Although no decreases in receptor level were observed at day 8, significant increases in receptor level in all three brain regions were detected at days 14 and 21 post-lesion. The results demonstrate that midbrain dopaminergic innervation exerts tonic effects on the levels of dopamine D2 receptor and mRNA in the caudate-putamen and the nucleus accumbens of the rat. Changes in receptor level are frequently accompanied by comparable changes in mRNA level, indicating a mass action relationship between receptor level and receptor biosynthesis in these forebrain regions in the rat.

Involvement of Dopamine D1 and D2 Receptors in the Regulation of Proenkephalin mRNA Abundance in the Striatum and Accumbens of the Rat Brain

Abstract: The effects of short‐term treatment (6 h) with selective D1 or D2 agonists and antagonists on the mRNA for proenkephalin in the medial and anterior aspects of the cau‐date‐putamen and the nucleus accumbens were assessed by in situ hybridization histochemistry. Proenkephalin mRNA abundance was significantly changed in the striatum and accumbens in response to D2 receptor manipulation. D2 blockade with haloperidol or raclopride increased, whereas D2 stimulation with LY‐171555 (D2 agonist) decreased, striatal and accumbens proenkephalin mRNA abundance. Antagonism of D1, receptor activity with SCH‐23390 significantly decreased proenkephalin mRNA abundance in all brain regions. Concurrent administration of the D1, agonist SKF‐38393 prevented the SCH‐23390 effect in all brain areas. The data demonstrate that acute treatment with dopaminergic D2 agonists and antagonists affects proenkephalin mRNA abundance in the striatum and accumbens via a D2 receptor mechanism, consistent with the concept that D2 receptor function inhibits the synthesis of the mRNA encoding the enkephalin peptides. Moreover, Df receptor activity, directly or indirectly, exerts modulatory effects on proenkephalin mRNA abundance in the striatum and nucleus accumbens.

Effect of chronic typical and atypical neuroleptic treatment on proenkephalin mRNA levels in the striatum and nucleus accumbens of the rat

Abstract: We measured proenkephalin (PEK) mRNA levels in the anterior and medial aspects of the caudate‐putamen (CPU) and in the nucleus accumbens (NAc) of the rat by in situ hybridization histochemistry after chronic treatment for 21 days with typical (haloperidol and prolixin) and atypical (molindone, thioridazine, and clozapine) neuroleptics. Chronic administration with these drugs resulted in PEK mRNA levels that were 60–80% higher than controls in the anterior and medial aspects of the CPU but only 25–30% over controls in the NAc. All three atypical neuroleptics studied increased PEK mRNA in the following order: anterior‐CPU, thioridazine > clozapine and molindone; medial‐CPU, thioridazine and molindone > clozapine; and NAc, thioridazine > > molindone and clozapine. Chronic treatment with the specific dopamine D2 antagonist sulpiride also caused elevation in PEK mRNA levels in all three brain regions studied whereas the specific serotonin S2 receptor blocker, cinanserin, had no significant effects on PEK mRNA levels. These results are consistent with the hypothesis that elevated levels of the enkephalins in the mesolimbic system may be necessary for antipsychotic activity. They also support the idea that the undesirable motoric signs and symptoms observed after chronic treatment with typical neuroleptics may not be the result of increased levels of enkephalins in the basal ganglia because atypical neuroleptics which are almost totally devoid of these side effects caused similar increases in PEK mRNA in the CPU.

Isolation stress increases tyrosine hydroxylase mRNA in the locus coeruleus and midbrain and decreases proenkephalin mRNA in the striatum and nucleus accumbens

Isolation of adult animals represents a form of psychological stress from which the animals cannot escape. In order to assess the effect of this stressor on neurochemical substrates in the brain, we assessed behavior and measured tyrosine hydroxylase and proenkephalin mRNA levels in selected brain areas by in situ hybridization histochemistry. Tyrosine hydroxylase (TH) mRNA levels in the locus coeruleus (LC) were significantly and progressively increased by 18, 42 and 68% after 7, 14 or 28 days of isolation, respectively. TH mRNA in the midbrain was transiently increased by isolation. Levels were significantly elevated by 34 and 48% above group-housed controls in the ventral tegmentum and the substantia nigra, respectively, after 14 days of isolation. In the forebrain, proenkephalin (PE) mRNA levels were found to be transiently decreased by 29% in the anterior and medial aspects of the caudate-putamen and the nucleus accumbens after 7 or 14 days of isolation stress, but the levels returned toward control levels after 28 days of isolation. Behavioral tests indicate that isolated animals progressively became more aggressive with duration of stress and showed a small but significant decrease in locomotor activity. The results demonstrate that a physically noninvasive stressor such as isolation of adult male rats can produce significant alterations in brain neurochemistry. The neurochemical responses observed may represent a brain mechanism designed to help the organism adapt to or protect from the deleterious effects of chronic psychological stress.

Disparity in the temporal appearance of methamphetamine-induced apoptosis and depletion of dopamine terminal markers in the striatum of mice

Methamphetamine (METH) causes damage in the striatum at pre- and post-synaptic sites. Exposure to METH induces long-term depletions of dopamine (DA) terminal markers such as tyrosine hydroxylase (TH) and DA transporters (DAT). METH also induces neuronal apoptosis in some striatal neurons. The purpose of this study is to demonstrate which occurs first, apoptosis of some striatal neurons or DA terminal toxicity in mice. This is important because the death of striatal neurons leaves the terminals in a state of deafferentation. A bolus injection (i.p.) of METH (30 mg/kg) induces apoptosis (TUNEL staining) in approximately 25% of neurons in the striatum at 24 h after METH. However, in contrast to apoptosis, depletion of TH (Western blotting) begins to appear at 24 h after METH in dorsal striatum while the ventral striatum is unaffected. The peak of TH depletion (approximately 80% decrease relative to control) occurs at 48 h after METH. Autoradiographic analysis of DAT sites showed that depletion begins to appear 24 h after METH and peaks at 2 days (approximately 60% depletion relative to control). Histological analysis of the induction of glial fibrillary acidic protein (GFAP) by METH in striatal astrocytes revealed an increase at 48 h after METH that peaked at 3 days. These data demonstrate that striatal apoptosis precedes the depletion (toxicity) of DA terminal markers in the striatum of mice, suggesting that the ensuing state of deafferentation of the DA terminals may contribute to their degeneration.

Increase in striatal dopamine D2 receptor mRNA after lesions of haloperidol treatment

In the present study, we show that both destruction of nigral dopamine neurons by 6-hydroxydopamine (6-OHDA) and chronic treatment with haloperidol increase the content of D 2 mRNA and D 2 receptors within distinct parts of the striatum

Quantitative in situ hybridization evidence for differential regulation of proenkephalin and dopamine D2 receptor mRNA levels in the rat striatum: effects of unilateral intrastriatal injections of 6-hydroxydopamine

Nigrostriatal (NS) dopaminergic (DA) neurons are thought to exert an inhibitory influence on striatal enkephalinergic systems through their DA D2 receptors. In order to investigate the effects of partial lesions of the NS DA on striatal proenkephalin (PEK) and D2 receptor mRNAs, animals were allocated to High, Intermediate, and Low rotators on the basis of amphetamine-induced rotation observed after intrastriatal injections of 6-hydroxydopamine (6-OHDA). On the ipsilateral side of the lesions, there were significant increases in PEK mRNA in the total aspect of the caudate-putamen (CPu) of the High (+204%), the Intermediate (+125%), and of the Low (+67%) rotation groups in comparison to controls; these changes correlated positively with increases in rotation rate. Unexpectedly, there were also significant increases in striatal PEK mRNA on the contralateral side although these changes were much less prominent than those observed on the side of the lesions. Conversely, only the High rotation group showed significant increases (+112%) in D2 receptor mRNA which occurred only on the lesioned side. Interestingly, the low rotation group actually showed some non-significant decreases (-25%) on the side of the lesions. These results indicate that partial lesions of the NS DA projections are sufficient to cause substantial increases in PEK mRNA but not in D2 receptor mRNA. These data also provide evidence that the two nigrostriatal DA projections and the systems which they modulate might be under interdependent sets of controls.(ABSTRACT TRUNCATED AT 250 WORDS)