Social dominance in monkeys: Lack of effect on ethanol self-administration during schedule induction.

Nonhuman primate models of alcohol use disorder (AUD) frequently utilize schedule-induced polydipsia to initiate ethanol drinking. Previous research has demonstrated that specific characteristics of drinking during the final phase of induction, in which monkeys consume 1.5 g/kg of ethanol per day, can predict whether monkeys become heavy or light drinkers when they subsequently have free access to ethanol (22 hours per day; Baker, Farro, Gonzales, Helms, & Grant, 2017; Grant et al., 2008). A monkey's position in the social dominance hierarchy is another factor associated with ethanol drinking in nonhuman primates; lower social status is associated with higher ethanol intakes. In the present study, characteristics of drinking during induction were measured in 12 male cynomolgus monkeys living in three established social groups (4 monkeys per group). All monkeys were induced to consume water, then increasing doses of ethanol (0.5, 1.0, and 1.5 g/kg) for 30 sessions per dose using a 300-s fixed-time schedule of food pellet delivery. Drinking sessions occurred five days per week and monkeys were group-housed on the other two days. Contrary to our hypothesis that subordinate monkeys would show characteristics of drinking during the last phase of induction that were predictive of later heavy drinking, no significant differences were observed between dominant and subordinate monkeys in any phase of induction. When ethanol availability was subsequently increased to 22 hours per day for 5 weeks, the intakes of subordinate- and dominant-ranked monkeys diverged, with higher intakes on average in subordinates. Several factors unique to the conditions of induction may have obscured any influence of social rank, including the limited duration of sessions and limited maximal ethanol intake. The data support the conclusion that the effects of social rank on ethanol consumption require unrestricted access to ethanol.

Functional connectivity in frontal-striatal brain networks and cocaine self-administration in female rhesus monkeys.

RATIONALE: Cocaine addiction is characterized by alternating cycles of abstinence and relapse and loss of control of drug use despite severe negative life consequences associated with its abuse. OBJECTIVE: The objective of the present study was to elucidate critical neural circuits involved in individual vulnerabilities to resumption of cocaine self-administration following prolonged abstinence. METHODS: The subjects were three female rhesus monkeys in prolonged abstinence following a long history of cocaine self-administration. Initial experiments examined the effects of acute cocaine administration (0.3 mg/kg, IV) on functional brain connectivity across the whole brain and in specific brain networks related to behavioral control using functional magnetic resonance imaging in fully conscious subjects. Subsequently, these subjects were allowed to resume cocaine self-administration to determine whether loss of basal connectivity within specific brain networks predicted the magnitude of resumption of cocaine intake following prolonged abstinence. RESULTS: Acute cocaine administration robustly decreased global functional connectivity and selectively impaired top-down prefrontal circuits that control behavior, while sparing connectivity of striatal areas within limbic circuits. Importantly, impaired connectivity between prefrontal and striatal areas during abstinence predicted cocaine intake when these subjects were provided renewed access to cocaine. CONCLUSIONS: Based on these findings, loss of prefrontal to striatal functional connectivity may be a critical mechanism underlying the negative downward spiral of cycles of abstinence and relapse that characterizes cocaine addiction.

Disruptions in serotonergic regulation of cortical glutamate release in primate insular cortex in response to chronic ethanol and nursery rearing.

Early-life stress has been shown to increase susceptibility to anxiety and substance abuse. Disrupted activity within the anterior insular cortex (AIC) has been shown to play a role in both of these disorders. Altered serotonergic processing is implicated in controlling the activity levels of the associated cognitive networks. We therefore investigated changes in both serotonin receptor expression and glutamatergic synaptic activity in the AIC of alcohol-drinking rhesus monkeys. We studied tissues from male rhesus monkeys raised under two conditions: Male rhesus monkeys (1) "mother reared" (MR) by adult females (n=9) or (2) "Nursery reared" (NR), that is, separated from their mothers and reared as a separate group under surrogate/peer-reared conditions (n=9). The NR condition represents a long-standing and well-validated nonhuman primate model of early life stress. All monkeys were trained to self-administer ethanol (4% w/v) or an isocaloric maltose-dextrin control solution. Subsets from each rearing condition were then given daily access to ethanol, water, or maltose-dextrin for 12 months. Tissues were collected at necropsy and were further analyzed. Using real time RT-PCR we found that ethanol-naive, NR monkeys had lower AIC levels of 5-HT(1A) and 5-HT(2A) receptor mRNA compared with ethanol-naive, MR animals. Although NR monkeys consumed more ethanol over the 12-month period compared with MR animals, both MR and NR animals expressed greater 5-HT(1A) and 5-HT(2A) receptor mRNA levels following chronic alcohol self-administration. The interaction between nursery-rearing conditions and alcohol consumption resulted in a significant enhancement of both 5-HT(1A) and 5-HT(2A) receptor mRNA levels such that lower expression levels observed in nursery-rearing conditions were not found in the alcohol self-administration group. Using voltage clamp recordings in the whole cell configuration we recorded excitatory postsynaptic currents in both ethanol-naive and chronic self-administration groups of NR and MR monkeys. Both groups that self-administered ethanol showed greater glutamatergic activity within the AIC. This AIC hyperactivity in MR alcohol-consuming monkeys was accompanied by an increased sensitivity to regulation by presynaptic 5-HT(1A) receptors that was not apparent in the ethanol-naive, MR group. Our data indicate that chronic alcohol consumption leads to greater AIC activity and may indicate a compensatory upregulation of presynaptic 5-HT(1A) receptors. Our results also indicate that AIC activity may be less effectively regulated by 5-HT in ethanol-naive NR animals than in NR monkeys in response to chronic ethanol self-administration. These data suggest possible mechanisms for increased alcohol seeking and possible addiction potential among young adults who had previously experienced early-life stress that include disruptions in both AIC activity and serotonin system dynamics.

The native T-type calcium current in relay neurons of the primate thalamus.

The generation of thalamic bursts depends upon calcium currents that flow through transiently open (T)-type calcium channels. In this study, we characterized the native T-type calcium current underlying thalamic burst responses in the macaque monkey. Current clamp recordings from lateral geniculate nucleus (LGN) slices showed characteristic burst responses when relay cells were depolarized from relatively hyperpolarized membrane potentials. These bursts could also be elicited by stimulation of excitatory synaptic inputs to LGN cells. Under voltage clamp conditions, the inactivation kinetics of native currents recorded from primate LGN neurons showed consistency with T-type currents recorded in other mammals and in expression systems. Real-time reverse transcriptase PCR performed on RNA isolated from the LGN (including tissues isolated from magnocellular and parvocellular laminae) detected voltage-dependent calcium channel (Ca(v)) 3.1, Ca(v) 3.2, and Ca(v) 3.3 channel transcripts. Ca(v) 3.1 occurred at relatively higher expression than other isoforms, consistent with in situ hybridization studies in rats, indicating that the molecular basis for burst firing in thalamocortical systems is an important conserved property of primate physiology. Since thalamic bursts have been observed during visual processing as well as in a number of CNS disorders, studies of the expression and modulation of these currents at multiple levels are critical for understanding their role in vision and for the discovery of new treatments for disruptions of thalamic rhythms.

Ethanol modulation of excitatory and inhibitory synaptic transmission in rat and monkey dentate granule neurons.

BACKGROUND: The physiological mechanisms underlying the behavioral and cognitive effects of ethanol are not fully understood. However, there is now compelling evidence that ethanol acts, at least in part, by modulating the function of a small group of proteins that mediate excitatory and inhibitory synaptic transmission. For example, intoxicating concentrations of ethanol have been shown to enhance GABAergic synaptic inhibition and depress glutamatergic excitatory neurotransmission in a number of brain regions. Because all of these electrophysiological studies have been performed in rodent brain slice or neuronal culture preparations, direct evidence that ethanol exerts similar effects on synaptic transmission in the primate central nervous system is lacking. METHODS: We have therefore developed methods to perform patch-clamp electrophysiological recordings from neurons in acutely prepared monkey (Macaca fascicularis) hippocampal slices. We have used these methods to compare the acute effects of ethanol on excitatory and inhibitory synaptic transmission in rat and monkey dentate granule neurons. RESULTS: Under our recording conditions, ethanol significantly potentiated gamma-aminobutyric acid type A inhibitory postsynaptic currents in both rat and monkey neurons. In addition, ethanol significantly inhibited NMDA, but not AMPA, excitatory postsynaptic currents in dentate granule neurons from both species. Notably, no significant differences were observed in any of the pharmacological properties of inhibitory or excitatory synaptic responses recorded from rat and monkey neurons. CONCLUSIONS: These data suggest that the differences in the behavioral effects of ethanol that have been observed between rats and higher-order mammals, such as monkeys and humans, may not reflect differences in the sensitivity of some of the major synaptic sites of ethanol action. Moreover, our results provide empirical evidence for the use of rodent brain slice preparations in elucidating synaptic mechanisms of ethanol action in the primate central nervous system.

Temporal upregulation of prodynorphin mRNA in the primate striatum after cocaine self-administration.

Several human and rat studies suggest that the striatal dynorphin system is important for neuroadaptation following cocaine exposure. In the current study, prodynorphin (PDYN) mRNA expression was examined in monkeys at initial and chronic phases of cocaine self-administration. Adult Rhesus monkeys were trained to self-administer food (banana flavoured pellets) or cocaine (0.03 or 0.3 mg/kg per injection) on a fixed interval 3-min schedule for 5 or 100 sessions. Each session ended after 30 reinforcers were delivered. The PDYN mRNA expression was analysed in the precommissural striatum using in situ hybridization histochemistry. We found a specific activation of PDYN mRNA expression in the limbic-innervated patch/striosome compartment of the dorsal caudate and dorsal putamen during the initial (i.e. 5 day) phase of the high dose cocaine self-administration. After 100 days of the high dose exposure, the patch/striosome compartment remained activated, but an increase in PDYN mRNA levels was also evident in the sensorimotor-connected matrix compartment of the caudate. Neither self-administration phase resulted in significant changes in the corresponding striatal regions of the low dose cocaine-exposed primates. Moreover, cocaine self-administration failed to alter the PDYN mRNA expression in high- or low-expressing PDYN cell populations in the nucleus accumbens during any condition studied. These results demonstrate the vulnerability of the dorsal striatum (in particular the caudate) to neuroadaptations following long-term high dose cocaine self-administration. In addition, the temporal nature of the changes in PDYN gene expression within the striatal compartments could reflect a change in drug responsivity that occurs during the transition to drug dependence.

Functional and anatomical localization of mu opioid receptors in the striatum, amygdala, and extended amygdala of the nonhuman primate.

The subregional distribution of mu opioid receptors and corresponding G-protein activation were examined in the striatum, amygdala, and extended amygdala of cynomolgus monkeys. The topography of mu binding sites was defined using autoradiography with [(3)H]DAMGO, a selective mu ligand. In adjacent sections, the distribution of receptor-activated G proteins was identified with DAMGO-stimulated guanylyl 5'(gamma-[(35)S]thio)triphosphate ([(35)S]GTPgammaS) binding. Within the striatum, the distribution of [(3)H]DAMGO binding sites was characterized by a distinct dorsal-ventral gradient with a higher concentration of binding sites at more rostral levels of the striatum. [(3)H]DAMGO binding was further distinguished by the presence of patch-like aggregations within the caudate, as well as smaller areas of very dense receptor binding sites, previously identified in human striatum as neurochemically unique domains of the accumbens and putamen (NUDAPs). The amygdala contained the highest concentration of [(3)H]DAMGO binding sites measured in this study, with the densest levels of binding noted within the basal, accessory basal, paralaminar, and medial nuclei. In the striatum and amygdala, the distribution of DAMGO-stimulated G-protein activation largely corresponded with the distribution of mu binding sites. The central and medial nuclei of the amygdala, however, were notable exceptions. Whereas the concentration of [(3)H]DAMGO binding sites in the central nucleus of the amygdala was very low, the concentration of DAMGO-stimulated G-protein activation in this nucleus, as measured with [(35)S]GTPgammaS binding, was relatively high compared to other portions of the amygdala containing much higher concentrations of [(3)H]DAMGO binding sites. The converse was true in the medial nucleus, where high concentrations of binding sites were associated with lower levels of DAMGO-stimulated G-protein activation. Finally, [(3)H]DAMGO and [(35)S]GTPgammaS binding within the amygdala, particularly the medial nucleus, formed a continuum with the substantia innominata and bed nucleus of the stria terminalis, supporting the concept of the extended amygdala in primates.

Chronic cocaine-mediated changes in non-human primate nucleus accumbens gene expression.

Chronic cocaine use elicits changes in the pattern of gene expression within reinforcement-related, dopaminergic regions. cDNA hybridization arrays were used to illuminate cocaine-regulated genes in the nucleus accumbens (NAcc) of non-human primates (Macaca fascicularis; cynomolgus macaque), treated daily with escalating doses of cocaine over one year. Changes seen in mRNA levels by hybridization array analysis were confirmed at the level of protein (via specific immunoblots). Significantly up-regulated genes included: protein kinase A alpha catalytic subunit (PKA(calpha)); cell adhesion tyrosine kinase beta (PYK2); mitogen activated protein kinase kinase 1 (MEK1); and beta-catenin. While some of these changes exist in previously described cocaine-responsive models, others are novel to any model of cocaine use. All of these adaptive responses coexist within a signaling scheme that could account for known inductions of genes(e.g. fos and jun proteins, and cyclic AMP response element binding protein) previously shown to be relevant to cocaine's behavioral actions. The complete data set from this experiment has been posted to the newly created Drug and Alcohol Abuse Array Data Consortium (http://www.arraydata.org) for mining by the general research community.

A cocaine analog, 2beta-propanoyl-3beta-(4-tolyl)-tropane (PTT), reduces tyrosine hydroxylase in the mesolimbic dopamine pathway.

Tyrosine hydroxylase (TH) is the rate-limiting enzyme in catecholamine biosynthesis. Previously published results have established that chronic cocaine administration (30-45 mg/kg per day, 10-14 days) resulted in an upregulation of TH gene expression in dopaminergic pathways of rats. The present studies tested the effects of a tropane analog, PTT (2beta-propanoyl-3beta-(4-tolyl)-tropane), on TH expression. This drug has similar actions to cocaine, but possesses markedly different pharmacokinetics (20 times more potent at binding the dopamine transporter, markedly increased metabolic stability, and 10-20 times more potent in behavioral measures). Moreover, PTT demonstrates an increased selectivity for the dopamine (DA) and norepinephrine (NE) transporters compared with cocaine. In direct contrast to the previously reported effects of cocaine, 10 days of PTT administration (3.0 mg/kg per day, i.p.) produced a uniform downregulation of TH protein and activity gene expression. TH activity and immunoreactive protein where decreased by 54 and 69%, respectively in the nucleus accumbens. Within the ventral tegmental area, TH activity and protein were decreased by 33 and 19%, respectively. The underlying mechanisms for these fundamental differences are unclear, but likely reflect varying and selective affinities and lengths of occupancy at biogenic amine transporters.

A comparison of the behavioral effects of the repeated administration of PTT, 2beta-propanoyl-3beta-(4-tolyl)tropane and cocaine.

In the present study the effects of the repeated administration of the novel tropane analog PTT (2beta-propanoyl-3beta(4-tolyl)tropane) on spontaneous locomotor activity were compared to those of cocaine. Previous reports describing the in vivo effects of PTT have focused solely on its acute effects following a single administration. In Experiment 1, PTT (1.0, 3.0 mg/kg), cocaine (30 mg/kg), or vehicle were administered intraperitoneally to male Sprague-Dawley rats daily for 10 consecutive days and locomotor activity was assessed. In Experiment 2, the locomotor effects of PTT (1.0 mg/kg) and cocaine (15 mg/kg) were assessed following 5 days of drug exposure to either PTT (1.0, 3.0 mg/kg) or cocaine (30 mg/kg) and 18 days of withdrawal. In both paradigms, PTT (1.0, 3.0 mg/kg) and cocaine (30 mg/kg) produced marked increases in locomotor activity acutely and an augmented response to drug challenge following repeated exposure, indicative of behavioral sensitization. These data indicate that, despite differences in the pharmacological profiles of PTT and cocaine, both drugs produce behavioral sensitization. These data are consistent with previous reports in the literature describing the effects of the repeated administration of other dopamine reuptake inhibitors and suggest that the development of behavioral sensitization is a uniform characteristic of this class of drugs.

Mu and kappa1 opioid-stimulated [35S]guanylyl-5'-O-(gamma-thio)-triphosphate binding in cynomolgus monkey brain.

Agonist-stimulated [35S]GTPgammaS binding allows the visualization of receptor-activated G-proteins, thus revealing the anatomical localization of functional receptor activity. In the present study, agonist-stimulated [35S]GTPgammaS binding was used to demonstrate mu and kappa1 opioid-stimulated [35S]GTPgammaS binding in tissue sections and membranes from cynomolgus monkey brain using DAMGO and U50,488H, respectively. Concentrations of agonists required to produce maximal stimulation of [35S]GTPgammaS binding were determined in membranes from the frontal poles of the brain. Receptor specificity was verified in both membranes and sections by inhibiting agonist-stimulated [35S]GTPgammaS binding with the appropriate antagonist. Mu opioid-stimulated [35S]GTPgammaS binding was high in areas including the amygdala, ventral striatum, caudate, putamen, medial thalamus and hypothalamus. Dense mu-stimulated [35S]GTPgammaS binding was also found in brainstem nuclei including the interpeduncular nucleus, parabrachial nucleus and nucleus of the solitary tract. Kappa1 opioid-stimulated [35S]GTPgammaS binding was high in limbic and association cortex, ventral striatum, caudate, putamen, globus pallidus, claustrum, amygdala, hypothalamus and substantia nigra. These results demonstrate the applicability of [35S]GTPgammaS autoradiography to examine receptor-activated G-proteins in the primate brain and reveal functional mu and kappa1 opioid receptor activity that may contribute to the reported central nervous system effects of opiates.

Long-acting blockade of biogenic amine transporters in rat brain by administration of the potent novel tropane 2beta-propanoyl-3beta-(2-Naphthyl)-tropane.

2beta-Propanoyl-3beta-(2-naphthyl)-tropane (WF-23) is a potent cocaine analog with activity at dopamine and serotonin transporters. The purpose of these experiments was to characterize the time course of effects of acute administration of WF-23 on spontaneous locomotion and biogenic amine transporters. Rats received injections i.p. with WF-23 (1 mg/kg), cocaine (30 mg/kg) or vehicle and locomotor activity was measured at various times postinjection. Animals were killed immediately after behavioral activity. Locomotor activity was significantly increased by WF-23 administration, reaching maximum at 4 hr and persisting for 24 hr. Cocaine-elicited elevations in locomotor activity occurred only at the earliest times. WF-23 decreased DA transporter binding in striatal membranes ([125I]RTI-55 binding), with >50% loss in binding for up to 49 hr postinjection. WF-23 increased the Kd of the high affinity site, with no effect on Bmax. Cocaine depressed binding (20%) only at the earliest times. WF-23 decreased levels of [3H]WIN 35,428 binding sites up to 95% of control in both dorsal and ventral striatum with a similar time-course when assessed autoradiographically. WF-23 also reduced [3H]citalopram binding to serotonin transporter sites throughout the brain. The slow onset and very long duration of action of WF-23, taken together with its actions at dopamine and serotonin transporters, suggest a potential role for treatment of disorders characterized by their involvement of these neural systems.

Alterations in behavior and opioid gene expression induced by the novel tropane analog WF-31.

The effects of the acute administration of the serotonin-selective tropane analog, [2beta-propanoyl-3beta-(4-isopropylphenyl)-tropane, WF-31, on spontaneous locomotor activity were measured and compared to those of the highly selective serotonin uptake inhibitor, fluoxetine and cocaine, a non-selective re-uptake inhibitor of dopamine and serotonin. WF-31 (1, 10 and 30 mg/kg)-elicited increases in locomotor behaviors when compared to vehicle-treated rats. This increased activity was blocked by pre-treatment with the dopaminergic antagonist, flupenthixol, suggesting that these effects may be mediated by dopaminergic mechanisms. Cocaine, but not fluoxetine, also elicited increases in behaviors. In addition, the effects of these three compounds on opioid peptide gene expression were also assessed using in situ hybridization histochemistry in the same animals. The acute administration of both WF-31 and cocaine increased the expression of preprodynorphin mRNA in the dorsal striatum whereas fluoxetine had no effect. Expression of striatal preproenkephalin mRNA was augmented by all three compounds. Within the nucleus accumbens, PPD mRNA levels were affected only by treatment with WF-31, an effect that was blocked by pre-treatment with flupenthixol. In contrast, the acute administration of both WF-31 and fluoxetine, but not cocaine, increased the expression of preproenkephalin mRNA. These increases, however, were not reversed by pre-treatment with flupenthixol. Despite its profile in vitro as a relatively selective serotonin re-uptake inhibitor, some of the in vivo actions of WF-31 appear to be mediated by dopaminergic mechanisms. These data further suggest that the mechanisms underlying expression of the opioid peptides in the nucleus accumbens may vary from those in the dorsal striatum.

Long-term cocaine self-administration decreases striatal preproenkephalin mRNA in rhesus monkeys.

The consequences of long-term exposure to cocaine on striatal preproenkephalin mRNA were assessed using quantitative in situ hybridization in monkeys that self-administered cocaine for approximately 2 years. Autoradiograms revealed high levels of preproenkephalin hybridization signal in both the caudate and putamen of the dorsal striatum, as well as in the shell and core subdivisions of the nucleus accumbens of drug-naive control monkeys. In general, there was a medial to lateral and ventral to dorsal gradient of preproenkephalin mRNA observed within the striatum of normal controls. Preproenkephalin mRNA was significantly reduced in widespread portions of both the dorsal and ventral striatum following chronic long-term cocaine self-administration when compared with levels in normal controls. These data confirm those observed in human drug abusers and suggest that long-term abuse of cocaine can result in significant alterations in the opioid regulation of striatal efferent neurons.

Effects of chronic cocaine administration on dopamine transporter mRNA and protein in the rat.

Male Sprague-Dawley rats were administered cocaine (10, 15 or 25 mg/kg) or vehicle, i.p., once daily for 8 consecutive days and killed 1 h after the last injection. Acute cocaine administration produced dose-dependent increases in spontaneous locomotor activity. These levels of activity were further enhanced by 8 days of chronic treatment, indicating the emergence of behavioral sensitization. Chronic cocaine administration resulted in dose-dependent decreases in the density of dopamine transporter (DAT) mRNA in both the substantia nigra pars compacta and ventral tegmental area as shown by in situ hybridization histochemistry. Changes in DAT binding sites were assessed using [3H]mazindol quantitative autoradiography. In contrast to the levels of mRNA, there were few changes in the number of [3H]mazindol binding sites. Although the density of binding sites was unaltered in most regions, [3H]mazindol binding was increased in the anterior nucleus accumbens. This study extends previous findings by demonstrating the dose-dependent nature of the changes in DAT mRNA that accompanies chronic cocaine administration. The levels of DAT binding sites within the dorsal and ventral striatum, however, were largely unchanged. This mismatch suggests that cocaine may differentially influence the gene expression of DAT in the ventral midbrain as compared to the density of DAT binding sites in the basal forebrain.

The effects of D1 or D2 dopamine receptor blockade on zif/268 and preprodynorphin gene expression in rat forebrain following a short-term cocaine binge.

Selective D1 or D2 dopamine receptor antagonists were used to investigate the transynaptic regulation of mRNAs coding for the opioid peptide, preprodynorphin, and the nuclear transcription factor, zif/268 after an acute cocaine binge. Rats were injected intraperitoneally with the D1 receptor antagonist, SCH 23390, or the D2 receptor antagonist, sulpiride, 30 min prior to 3 hourly injections of saline or 20 mg/kg cocaine and killed 1 h after the final injection. Behavioral ratings indicated that SCH 23390 blocked, whereas sulpiride augmented, cocaine-induced stereotypical behaviors. Striatal sections were hybridized with oligonucleotides coding for zif/268 and preprodynorphin. Quantitative image analysis of autoradiograms revealed that (1) SCH 23390 completely suppressed basal and cocaine binge-induced zif/268 mRNA in the striatal and cerebral cortical areas examined; (2) sulpiride enhanced basal levels of zif/268 mRNA in the medial caudate and dorsomedial shell of the nucleus accumbens; (3) sulpiride partially blocked cocaine binge-induced levels of zif/268 mRNA in the dorsal striatum but had no effect in sensory cortex; (4) SCH 23390, but not sulpiride, significantly reduced the constitutive expression of preprodynorphin mRNA; and (5) SCH 23390 and sulpiride blocked cocaine binge-induced expression of preprodynorphin mRNA in the dorsal striatum.

Cocaine binges differentially alter striatal preprodynorphin and zif/268 mRNAs.

Several studies have demonstrated that cocaine increases preprodynorphin, c-fos, and zif/268 mRNAs in rat dorsal striatum. Multiple, closely spaced exposures to cocaine appear to elicit the greatest increases in dynorphin. However, the response of preproenkephalin, c-fos and zif/268 mRNAs to such a dosing regimen is unknown. Therefore, we used a 'binge' paradigm to evaluate changes in mRNA for preprodynorphin, preproenkephalin, c-fos and zif/268. Male Wistar rats received three hourly i.p. injections of saline or 10 or 20 mg/kg cocaine for 1, 5, or 10 days. Although cocaine-induced locomotor and stereotypical behaviors were significantly increased as compared to saline on days 1, 5 and 10, these behaviors were significantly less on day 10 than on days 1 and 5. One hour after the last injection on days 1, 5, or 10, the rats were anesthetized and decapitated for quantitative in situ hybridization histochemistry. C-fos mRNA was undetectable in all treatment groups whereas zif/268 mRNA in the dorsal striatum was increased in a dose-dependent manner (20 mg/kg > 10 mg/kg) but the intensity of hybridization signal decreased over time (1 day >> 5 days > 10 days) as compared to that in saline-treated controls. In contrast, 10 mg/kg cocaine binges caused an increase in preprodynorphin, but not preproenkephalin, mRNA in the dorsal, but not ventral, striatum in a time-dependent manner (day 10 >> day 5 > day 1) whereas 20 mg/kg cocaine binges caused an increase in striatal preprodynorphin that was greater on day 1 and day 5 than on day 10. These data indicate that (1) c-fos, zif/268 and preprodynorphin mRNAs are differentially regulated in dorsal striatum, (2) behavioral tolerance results from chronic binges with 10 and 20 mg/kg cocaine and (3) the preprodynorphin genomic response exhibits tolerance to chronic high dose, but not low dose, cocaine binges.

Short-term cocaine self administration alters striatal gene expression.

Rats self-administered cocaine or received saline during 3 daily 5 h sessions and were euthanized 1 h after the final session. Quantitative in situ hybridization revealed that cocaine self-administration increased levels of preprodynorphin, but not preproenkephalin, c-fos, or zif/268 mRNAs in a patchy pattern in the dorsal striatum. These data demonstrate that the regulation of preprodynorphin gene expression is dissociable from that of c-fos and zif/268 in dorsal striatum following short-term cocaine self-administration.

NMDA receptors mediate amphetamine-induced upregulation of zif/268 and preprodynorphin mRNA expression in rat striatum.

The role of N-methyl-D-aspartate (NMDA) excitatory amino acid receptors in D-amphetamine (AMPH)-induced behavioral changes and increased expression of the nuclear transcription factors, c-fos and zif/268, and preprodynorphin (PPD) mRNA in various regions of rat forebrain was investigated with quantitative in situ hybridization histochemistry. Three hours after a single injection of AMPH (5 mg/kg, i.p.), the mRNA expression of zif/268, but not c-fos, in dorsal striatum (caudate nucleus) and cerebral cortex (sensorimotor cortex), and PPD mRNA in dorsal striatum, was upregulated. Pretreatment of rats with MK-801 (0.5 mg/kg, i.p.) attenuated AMPH-induced striatal and cortical expression of zif/268 mRNA and striatal expression of PPD mRNA, without affecting the behavioral alterations induced by AMPH. A similar, dose-dependent suppression of AMPH-induced zif/268 and PPD mRNA in striatum and cortex was also revealed after systemic administration of (+/-)-3-(2-carboxypiperazin-4-yl)-propyl-1-phosphonic acid (CPP) at doses of 5 and 10 mg/kg. CPP, only at the higher dose, slightly attenuated behavioral activity induced by AMPH. MK-801 and CPP (at higher dose) alone suppressed basal (constitutive) zif/268 mRNA levels in both striatum and cortex regions. No significant effect of either antagonist was found on constitutive expression of striatal PPD mRNA. These studies indicate that NMDA receptors mediate, at least in part, activation of zif/268 and PPD gene expression in striatum and sensorimotor cortex by a single injection of AMPH. Furthermore, NMDA receptor-mediated gene regulation more likely is involved in long-term neuronal plasticity to drug exposure than in acute drug effects since NMDA receptor antagonists had little or no effect on the acute behavioral actions of AMPH.