Body weight: reduction by long-term glycerol treatment.

Elevation of body glycerol concentration by multiple daily injections of glycerol was shown to lead to hypophagia and body weight loss followed by normal food intake and normal rate of body weight increase in rats. Termination of injections was followed by hyperphagia and an accelerated rate of growth. These findings suggest that the blood glycerol concentration plays an important role in the control of body weight and may be one signal by which the central nervous system monitors body lipid content.

Sustained intracerebroventricular infusion of brain fuels reduces body weight and food intake in rats.

Long-term infusion of glucose, beta-hydroxybutyrate, and glycerol into the third ventricle of the rat brain caused a stabilization of body weight at a lower than normal level. Among the glucose- and glycerol-treated animals this weight loss was caused in part by temporary hypophagia. Among the animals treated with beta-hydroxybutyrate the weight loss was unaccompanied by a reduction in food intake. The results are consistent with the view that the systems controlling food intake and body weight are sensitive to the availability of brain fuels. They are not consistent however, with the view that these control systems monitor calories independently of their source.

Forebrain lesions differentially affect drinking elicited by dipsogenic challenges and injections of muscimol into the median raphe nucleus.

Injections of muscimol into the median raphe nucleus (MR) elicit intense drinking in normally hydrated rats. To determine whether this response is dependent on forebrain systems mediating other aspects of water intake, the authors examined the effects of lesions of the subfornical organ (SFO), median preoptic nucleus (MnPO), lateral preoptic area (LPO), or lateral hypothalamus (LH) on the drinking. Lesions of the SFO or LH attenuated muscimol-elicited drinking, whereas lesions of the MnPO or LPO increased water intake after the treatment. All of the lesion groups showed a deficit in drinking to injections of polyethylene glycol and at least one of the doses of hypertonic saline. Only the SFO- and LH-lesioned groups showed a suppression of drinking to systemic injections of angiotensin II, suggesting that the drinking elicited by intra-MR injections of muscimol may involve changes in the central circuits mediating angiotensin-induced drinking.

Control of food intake by kainate/quisqualate receptors in the median raphe nucleus.

In previous studies we have reported that increases in food and water intake can be produced by microinjections of both NMDA selective and broad spectrum excitatory amino acid antagonists into the median raphe nucleus (MR). In the current experiments we examined the influence of kainate/quisqualate receptors in the MR on ingestive behavior. The consumption of food and water by deprived rats could be suppressed by intra-MR microinjections of the excitatory amino acid agonists kainic acid (5-10 ng in 0.5 microliters vehicle) and quisqualic acid (125-500 ng). Conversely, intra-raphe injections of the kainate/quisqualate receptor antagonists pBB-PZDA (1.25-2.5 micrograms) and GAMS (10-20 micrograms) elicited feeding in nondeprived animals. pBB-PZDA was more potent in eliciting ingestive behavior than was the selective NMDA antagonist 2-amino-6-phosphonohexanoic acid, suggesting that the effects of pBB-PZDA were not mediated through the NMDA receptor. The current findings suggest that ingestive behaviors are tonically inhibited by excitatory amino acids acting at kainate/quisqualate receptors in the vicinity of the MR.

Clonidine produces a conditioned place preference in rats.

The possibility that the alpha-adrenergic agonist clonidine can act as a reinforcing agent was investigated using the conditioned place preference paradigm. Using two different variants of this method we were able to demonstrate reinforcing properties of clonidine at doses of 200 and 400 micrograms/kg. These results are consistent with those obtained by other investigators using the self-administration technique, and support the view that adrenergic mechanisms may be involved in reinforcement.

Serotonin-1B agonists induce compartmentally organized striatal Fos expression in rats.

The 5HT1B agonist RU24969 (2.5-5.0 mg/kg) and anpirtoline (2.0 mg/kg) induced a striking increase in striatal Fos-like immunoreactivity in rats. In the rostral and dorsal regions of the striatum staining was dense and relatively homogeneous. In the ventral region of the striatum at more caudal levels, however, both drugs induced staining in patches which were in register with the opiate receptor rich striosomes. The effects of RU24969 could not be antagonized by the selective 5HT1A antagonist p-MPPI and little or no striatal Fos expression could be observed after injections of the selective 5HT1A agonist 8-OHDPAT or the selective 5HT3 antagonist MDL-72222.

Dissociation of hippocampal serotonin release and locomotor activity following pharmacological manipulations of the median raphe nucleus.

In vivo microdialysis was used to investigate the role of serotonin in the locomotor hyperactivity produced by injections of 8-hydroxy-2-(di-n-propylamino)tetralin (8-OHDPAT), muscimol and baclofen into the median raphe nucleus (MR) of unanesthetized rats. Intra-MR injections of the GABA(A) agonist muscimol (25 ng) resulted in a pronounced increase in locomotor activity which was accompanied by a 42% decrease in hippocampal serotonin release during the first hour following injection. Intra-MR injections of the GABA(B) agonist baclofen (125 ng) induced hyperactivity of a similar magnitude, but failed to affect hippocampal serotonin release. In contrast, the serotonin (5-HT1A) agonist 8-OHDPAT (5 microg) produced only a small effect on locomotor activity but reduced hippocampal serotonin output by 51%. These findings demonstrate that it is possible to dissociate the effects of intra-MR drug injections on locomotor activity and hippocampal 5-HT release and strongly support the view that nonserotonergic neurons in the paramedian tegmentum are importantly involved in the control of behavioral arousal.

Studies on the behavioral activation produced by stimulation of GABAB receptors in the median raphe nucleus.

Injections of the GABAB agonist baclofen into the median raphe nucleus (MR) resulted in marked hyperactivity and in increases in food and water intake by non-deprived animals. The locomotor effects of baclofen were stereospecific and could be antagonized by coinjection of the GABAB antagonist 2-hydroxysaclofen. Hyperactivity was produced by lower doses of baclofen, at shorter latencies, when the drug was injected into the MR than when it applied to the dorsal raphe nucleus (DR) or the ventral tegmental area (VTA). The locomotor response to intra-MR baclofen was unaltered in animals pretreated with the serotonin synthesis inhibitor p-chlorophenylalanine. Finally, intra-MR injections of baclofen produced a large increase in dopamine metabolism in the nucleus accumbens and striatum but failed to alter hippocampal or striatal serotonin metabolism. These findings suggest that baclofen may produce increases in activity and ingestive behavior as a result of an action on non-serotonergic cells in the MR.

Nicotine-induced behavioral sensitization is associated with extracellular dopamine release and expression of c-Fos in the striatum and nucleus accumbens of the rat.

It is well known that repeated injections of nicotine produce progressively larger increases in locomotor activity, an effect referred to as behavioral sensitization. This study was carried out to investigate the neural mechanisms underlying nicotine-induced behavioral sensitization using in vivo microdialysis and Fos-like immunohistochemistry (FLI). Rats were given repeated injections of saline or nicotine (0.4 mg/kg s.c., twice daily for 7 days) followed by one challenge injection on the 4th day after the last daily injection. Systemic challenge with nicotine produced a much larger increase in locomotor activity in nicotine-pretreated rats (659.1+/-94.9 counts/2 h) than in saline-pretreated rats (218.1+/-61 counts/2 h). A direct local challenge of nicotine (1 or 5 mM) via a microdialysis probe in the nucleus accumbens or striatum induced a much greater dose-dependent increase of dopamine (DA) output in nicotine-pretreated rats than in saline-pretreated rats. Furthermore, in parallel with the behavioral and biochemical data, systemic challenge with nicotine produced marked Fos-like immunohistochemistry in the nucleus accumbens and the striatum in the nicotine-pretreated rats. Taken together, this study demonstrates that behavioral sensitization is clearly associated with an increase in DA release and activation of Fos-like immunoreactive cells in the striatum and the nucleus accumbens produced by repeated nicotine treatment. Our results strongly suggest that the striatum and the nucleus accumbens may play a major role in nicotine-induced behavioral sensitization. The present results are discussed in terms of the development and expression of nicotine-induced behavioral sensitization.

Suppression of locomotor activity produced by acute injections of kainic acid into the median raphe nucleus.

Several lines of evidence suggest that cells within the immediate vicinity of the median raphe nucleus may exert an inhibitory influence on locomotor activity. If this theory were correct, one would expect that excitation of neurons within the median raphe would have a suppressive effect on behavior. This possibility was examined in the present study where low doses of the glutamate analogue kainic acid were injected into the median raphe nucleus of methylphenidate-pretreated rats. Our results indicate that these injections were able to antagonize, in a dose-dependent manner, both the horizontal locomotor activity and the nose poking induced by methylphenidate. These results provide further support for the existence of inhibitory median raphe influences on locomotion and suggest the possibility that endogenous excitatory amino acids may play a role in raphe functioning.

Fos-like immunoreactivity in the mamillary body and thalamus following injections of muscimol into the ventral tegmental nucleus of Gudden in the rat.

In many neurons, increased rates of firing are accompanied by expression of the proto-oncoprotein Fos. The current study examined Fos-like immunoreactivity in the mamillary body and the anterior thalamus following unilateral injections of the inhibitory GABA-A agonist muscimol into the ventral tegmental nucleus of Gudden (VTN). These injections resulted in a marked increase in Fos-like immunoreactivity ipsilaterally in both the medial mamillary nucleus and in its principle thalamic projection targets, the anteroventral and anteromedial thalamic nuclei. Since the projection from the VTN to the mamillary body has been shown to contain a substantial GABAergic component, these results are likely to reflect a disinhibition of mamillothalamic circuitry resulting from suppression of tonic inhibitory inputs arising in the VTN.

Comparative effects of scopolamine and quinpirole on the striatal fos expression induced by stimulation of D(1) dopamine receptors in the rat.

Treatment of intact rats with the full D(1) dopamine agonist A-77636 induced Fos-like immunoreactivity in the medial and, to a lesser extent, the lateral portions of the striatum. Pretreatment with the muscarinic antagonist scopolamine hydrobromide (1.5-6 mg/kg) potentiated the response to A-77636 and eliminated the mediolateral staining gradient seen after A-77636 alone. Similar effects were not produced by scopolamine methylbromide, which fails to cross the blood-brain barrier, demonstrating that the actions of scopolamine were centrally mediated. The effects of scopolamine were further compared to those of the D(2)-like dopamine agonist quinpirole using a factorial design in which subjects were pretreated with either scopolamine, quinpirole, or a combination of the two drugs before receiving A-77636. Pretreatment with either scopolamine or quinpirole increased staining in the lateral striatum, but the combination of the two drugs was no more effective than was quinpirole alone. Pretreatment with quinpirole, but not scopolamine, resulted in a markedly "patchy" pattern of staining and actually suppressed staining in the region between patches in the medial striatum. These findings demonstrate that there are both differences and similarities between the effects of scopolamine and quinpirole on D(1) agonist-induced Fos expression and suggest that although inhibition of cholinergic neurons may be one of the mechanisms through which the effects of quinpirole are produced, other factors must also contribute.

Evidence for GABAergic projections from the tegmental nuclei of Gudden to the mammillary body in the rat.

Immunochemical studies have demonstrated the presence of large numbers of cells immunoreactive for glutamic acid decarboxylase (GAD) within the dorsal and ventral tegmental nuclei of Gudden. Following injections of Fluoro-Gold into the medial mammillary nucleus, a substantial proportion of the retrogradely labeled neurons within the ventral tegmental nucleus displayed GAD-like immunoreactivity. Conversely, electrolytic or excitotoxic lesions of the ventral tegmental nucleus produced a large decrease in the number of fibers and terminals immunoreactive for GAD within the medial mammillary nucleus. In contrast, electrolytic lesions of the dorsal tegmental nucleus were found to produce a large decrease in GAD-like immunoreactivity which was restricted to the lateral mammillary nucleus. Control lesions placed caudal to the dorsal tegmental nucleus were without effect. These findings suggest that the dorsal and ventral nuclei send a substantial, topographically organized, GABAeregic input to the mammillary body.

Dopamine antagonists induce fos-like-immunoreactivity in the substantia nigra and entopeduncular nucleus of the rat.

Injections of the D2 receptor antagonists haloperidol (0.5-8 mg/kg) and metoclopramide (6.25-50 mg/kg) in rats resulted in a dose dependent induction of Fos-like-immunoreactivity in the rostral portion of the entopeduncular nucleus (EPN) and in the medial portion of the pars reticulata of the substantia nigra (SNpr). Nigral staining occurred exclusively in neurons which were not immunoreactive for tyrosine hydroxylase and could be antagonized by pretreatment with the anticholinergic drug scopolamine (3 mg/kg). Effects were much less pronounced following injections of the selective D1 antagonist SCH-23390 (2-8 mg/kg). No staining could be observed following administration of the 5HT3 antagonist MDL-72222 (10 mg/kg) or the 5HT1/5HT2 antagonist metergoline (5 mg/kg), suggesting that the effects observed with dopamine antagonists were not secondary to actions at serotonin receptors. These results are consistant with the hypothesis that blockade of dopamine receptors results in a disinhibition of cells within the SNpr and EPN and further suggest that examination of immediate-early gene expression may provide a useful tool for studying the extrastriatal circuit engaged by manipulations of dopaminergic transmission.

Interactive effects of stimulation of D1 and D2 dopamine receptors on Fos expression in the lateral habenula.

We have previously shown that systemic administration of non-selective dopamine agonists results in a pronounced expression of the proto-oncoprotein Fos within the lateral habenula. In the current study we examined the effects of selective D1 and D2 dopamine receptor agonists on habenular Fos expression. Rats were injected with various doses of the selective D2 agonist quinpirole (0, 0.62 or 2.5 mg/kg) either alone or in combination with various doses of the selective full D1 agonist A-77636 (0, 0.75 or 3.0 mg/kg). The selective agonists, by themselves, induced only small increases in Fos-like immunoreactivity within the lateral habenula, but combinations of the two drugs resulted in a very robust response. These findings indicate that D1 and D2 receptor agonists interact to induce Fos expression within the habenula and that the nature of this interaction differs from that reported in the striatum and the globus pallidus.

Quinpirole attenuates the striatal immediate early gene expression, but not the hyperactivity, induced by the serotonin agonist RU-24969.

Systemic administration of the mixed 5-HT(1A/1B) agonist RU-24969 has been shown to produce a dramatic increase in locomotor activity and to induce robust c-Fos expression in the rat striatum. Previous studies have also shown that pretreatment with the D2-like dopamine agonist quinpirole virtually abolishes RU-24969-induced striatal c-Fos expression. The present study was undertaken to determine whether the effects of RU-24969 on immediate early gene expression extend to the additional Fos family transcription factors FosB and Fra-2. Additionally, this study quantitatively examined the effect of quinpirole pretreatment on the ability of RU-24969 to induce both locomotor hyperactivity and striatal immediate early gene expression. RU-24969 alone produced elevations in locomotor activity and induced clear expression of c-Fos, FosB and Fra-2 throughout the entire striatal complex. Quinpirole pretreatment virtually abolished RU-24969-induced expression of all three transcription factors, but did not alter the elevated locomotor activity produced by RU-24969. These results demonstrate that the effects of RU-24969 on locomotor activity can be dissociated from its effects on immediate early gene expression within the striatum.

Haloperidol induces Fos expression in the globus pallidus and substantia nigra of cynomolgus monkeys.

Systemic injections of the dopamine antagonist haloperidol (0.1-2.5 mg/kg) induced a dose dependent increase in Fos-like immunoreactivity (FLI) in the internal segment of the globus pallidus (GPi) and in the substantia nigra (SN) of cynomolgus monkeys. These findings are consistent with models of basal ganglia organization which predict that blockade of dopamine receptors should result in a disinhibition of cells in these structures. In the GPi, labeling was most pronounced along the ventral, lateral and medial borders of the nucleus and none of the pallidal cells expressing FLI were immunopositive for choline acetyltransferase. In the SN, immunoreactive nuclei were concentrated in the pars reticulata and the majority of labeled nigral neurons did not display tyrosine hydroxylase-like immunoreactivity. A small number of cells displaying FLI were also observed in the external pallidal segment, but no labeling was seen in the subthalamic nucleus. These findings indicate that blockade of dopamine receptors induces a characteristic pattern of Fos expression in the primate brain which strongly resembles that previously reported in rodents.

Unilateral dopamine depletion paradoxically enhances amphetamine-induced Fos expression in basal ganglia output structures.

The ability of amphetamine to induce expression of the immediate early gene protein, Fos, was examined by immunocytochemistry in animals with unilateral 6-hydroxydopamine lesions of the nigrostriatal bundle. Amphetamine induced Fos expression in the globus pallidus (GP) on the intact side of the brain, but this response was greatly attenuated on the dopamine-depleted side. In contrast, amphetamine induced little Fos expression in the entopeduncular nucleus (EPN) and the substantia nigra pars reticulata (SNpr) on the intact side of the brain, but resulted in pronounced expression in these structures on the lesioned side. These findings demonstrate that unilateral dopamine depletion results in a pathophysiological state in which some responses to amphetamine are attenuated while others are paradoxically potentiated. One explanation of these effects is that amphetamine may indirectly activate excitatory inputs to the SNpr and the EPN on both sides of the brain. On the intact side, these effects would be opposed by the simultaneous activation of inhibitory pathways arising in the striatum and the GP, with the result that little Fos expression would be seen. On the dopamine-depleted side, however, engagement of these inhibitory pathways would be attenuated and the unopposed effects of the excitatory inputs mobilized by amphetamine would result in exaggerated Fos synthesis.

Quinpirole attenuates the striatal fos expression induced by escape behavior.

Several studies have shown that the D2-like dopamine receptor agonist quinpirole is able to markedly potentiate the striatal Fos expression induced by D1 agonists. The present study examined the effects of quinpirole on the striatal Fos-like immunoreactivity (FLI) induced by escape behavior. Male rats were pretreated with either saline or quinpirole (0.156, 0.625, 1.25 or 2.5 mg/kg) and 30 min later, placed in a shuttle box and required to crossover every 30 s in order to escape mild footshock. Animals were sacrificed 30 min following the completion of a 1-h block of escape trials and sections through the striatum were processed for FLI. Pretreatment with quinpirole produced a marked, dose-dependent, attenuation of escape-induced FLI in the striatum. These findings demonstrate that quinpirole affects the striatal Fos expression induced by shuttling in a very different fashion than it does that induced by D1 agonists, and further support the view that dopaminergic mechanisms play an important role in behaviorally induced striatal Fos expression.

Ascending dopaminergic projections from the dorsal raphe nucleus in the rat.

The projections of putative dopamine containing cells within the dorsal raphe nucleus (DR) were studied using a combination of tyrosine hydroxylase (TH) immunocytochemistry and fluorescent retrograde tracing. Substantial numbers of TH-immunoreactive cells in the DR were found to project to the nucleus accumbens. Progressively smaller numbers of cells were found to project to the lateral septum and medial prefrontal cortex. Very few TH-immunoreactive cells projected to the dorsal striatum, and none to the substantia nigra. TH-immunoreactive cells did not display serotonin-like immunoreactivity. These findings indicate that the projection pattern of TH-immunoreactive cells within the dorsal raphe more closely resembles that of dopaminergic cells within the ventral tegmental area (VTA) than that of serotonergic cells within the DR.