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Background: Inhibition of NMDA receptor function in brain is believed to be an important action of ethanol (EtOH). To investigate EtOH inhibition of NMDA receptor responses in vivo, the interaction of these agents in brain after different routes of administration were investigated by using transcription factor Fos protein expression to follow NMDA receptor activation and EtOH inhibition of this response., Methods: The induction of Fos-like immunoreactivity (Fos-LI) in 38 regions of the rat brain was measured 2 hr after treatment with NMDA, EtOH, or both. To determine the relative contribution of abdominal drug effects on Fos induction, rats received either intraperitoneal (ip) or intragastric (ig) EtOH and ip or intravenous (iv) NMDA. Rats received EtOH (2.5 g/kg ip or 4 g/kg ig) or vehicle 15 min before NMDA (125 mg/kg ip or 60 mg/kg iv) or vehicle., Results: For the 38 forebrain regions examined, ip and iv NMDA significantly induced Fos-LI in 13 and 32 regions, respectively. These effects occurred without elicitation of tonic-clonic seizure activity and were strong after iv NMDA in the frontal, prefrontal, and cingulate cortices, supraoptic nucleus, anterior lateral septum, and dentate gyrus. For EtOH, prominent Fos-LI induction was found in the central amygdala, dorsolateral bed nucleus of the stria terminalis, Edinger-Westphal nucleus, and paraventricular hypothalamus. Despite ip and ig EtOH induction of Fos-LI in these regions, the major effect of EtOH was to block NMDA-induced Fos-LI in 8 of 13 (ip) and 27 of 32 (ig) of the NMDA-positive regions, respectively, including retrosplenial, cingulate, and medial prefrontal cortices, central amygdala, and taenia tecta., Conclusions: These results provide new evidence for the regionally specific functional interactions of EtOH on NMDA receptors in vivo. Moreover, these results support efforts to identify brain region-specific targets for EtOH and EtOH-induced changes in gene expression.

This paper summarizes a conference held at the National Institute of Child Health and Human Development on December 6-7, 1999, on self-injurious behavior [SIB] in developmental disabilities. Twenty-six of the top researchers in the U.S. from this field representing 13 different disciplines discussed environmental mechanisms, epidemiology, behavioral and pharmacological intervention strategies, neurochemical substrates, genetic syndromes in which SIB is a prominent behavioral phenotype, neurobiological and neurodevelopmental factors affecting SIB in humans as well as a variety of animal models of SIB. Findings over the last decade, especially new discoveries since 1995, were emphasized. SIB is a rapidly growing area of scientific interest to both basic and applied researchers. In many respects it is a model for the study of gene-brain-behavior relationships in developmental disabilities.

The rat has proven to be a valuable preclinical model for characterizing effects of psychotrophic drugs and for identifying new psychotherapeutic agents in pharmacological screens. However, substantial differences have been described between the rat and human brain in regard to the neuroanatomical distribution of some drug and neurotransmitter receptor binding sites. To assess the utility of the rat as a model for the neuroanatomical topography of 5-HT1A and type 1 benzodiazepine (BDZ) receptors in humans, the distribution of binding sites for 3H-8-OH-DPAT (5-HT1A agonist) and 3H-zolpidem (type 1 BDZ agonist) was compared with autoradiography in select regions of the rat and human brain. Concordance in the binding patterns for the two ligands was observed in several brain regions for the two species. However, substantial differences were also found in the topography of binding sites for the ligands in the rat and human brain. High 3H-8-OH-DPAT binding was seen in the dorsal raphe nucleus and hippocampal formation in both the rat and human brain. However, species differences were observed in the relative distribution of ligand binding among hippocampal subregions. In the cerebral cortex, the laminar distribution of 3H-8-OH-DPAT binding sites was notably different for rats and humans. In humans, outer cortical layers were most densely labeled with 3H-8-OH-DPAT, whereas in the rat cortex, the highest binding was in the inner layers. A striking difference between rats and humans was observed for 3H-8-OH-DPAT binding in the lateral septal nucleus, which was densely labeled in the rat but weakly labeled in humans. Substantial differences between rats and humans were also observed for 3H-zolpidem binding. In the rat brain, high densities of binding sites were found in the medial septum, inferior colliculus, and substantia nigra reticulata. These regions showed very low 3H-zolpidem binding in the human brain. Intermediate binding was seen in the rat cerebral cortex, and low binding was found in the hippocampus. By contrast, in humans, cerebral cortical regions were the most densely labeled of all regions studied, and certain hippocampal subregions exhibited relatively high binding. The striking neuroanatomical differences in 3H-8-OH-DPAT and 3H-zolpidem binding observed between rats and humans suggest that different functional consequences may be produced within specific brain regions after administration of drugs that influence 5-HT1A and type 1 BZD receptors.

Subanesthetic doses of ketamine have been shown to exacerbate symptoms in schizophrenia and to induce positive, negative, and cognitive schizophrenic-like symptoms in normal subjects. The present investigation sought to define brain regions affected by subanesthetic doses of ketamine, using high resolution autoradiographic analysis of 14C-2-deoxyglucose (2-DG) uptake and immunocytochemical staining for Fos-like immunoreactivity (Fos-LI). Both functional mapping approaches were used because distinct and complementary information is often obtained with these two mapping methods. Ketamine, at a subanesthetic dose of 35 mg/kg, substantially increased 2-DG uptake in certain limbic cortical regions, including medial prefrontal, ventrolateral orbital, cingulate, and retrosplenial cortices. In the hippocampal formation, the subanesthetic dose of ketamine induced prominent increases in 2-DG uptake in the dentate gyrus, CA-3 stratum radiatum, stratum lacunosum moleculare, and presubiculum. Increased 2-DG uptake in response to 35 mg/kg ketamine was also observed in select thalamic nuclei and basolateral amygdala. Ketamine induced Fos-LI in the same limbic cortical regions that exhibited increased 2-DG uptake in response to the subanesthetic dose of the drug. However, no Fos was induced in some brain regions that showed increased 2-DG uptake, such as the hippocampal formation, anterioventral thalamic nucleus, and basolateral amygdala. Conversely, ketamine induced Fos in the paraventricular nucleus of the hypothalamus and central amygdala, although no effect of the drug on 2-DG uptake was apparent in these regions. In contrast to the increase in 2-DG uptake observed in select brain regions after the subanesthetic dose, an anesthetic dose of ketamine (100 mg/kg) produced a global suppression of 2-DG uptake. By contrast, a robust induction of Fos-LI was observed after the anesthetic dose of ketamine that was neuroanatomically identical to that produced by the subanesthetic dose. Results of the present investigation show that anesthetic and subanesthetic doses of ketamine have pronounced effects on regional brain 2-DG uptake and induction of Fos-LI. The alterations in regional brain metabolism induced by the subanesthetic dose may be relevant to effects of ketamine to induce schizophrenic-like symptoms., (Copyright 1998 Elsevier Science B.V.)

Dopamine (DA) deficiency has been implicated in Lesch-Nyhan disease (LND), a genetic disorder that is characterized by hyperuricemia, choreoathetosis, dystonia, and compulsive self-injury. To establish that DA deficiency is present in LND, the ligand WIN-35,428, which binds to DA transporters, was used to estimate the density of DA-containing neurons in the caudate and putamen of six patients with classic LND. Comparisons were made with 10 control subjects and 3 patients with Rett syndrome. Three methods were used to quantify the binding of the DA transporter so that its density could be estimated by a single dynamic positron emission tomography study. These approaches included the caudate- or putamen-to-cerebellum ratio of ligand at 80-90 min postinjection, kinetic analysis of the binding potential [Bmax/(Kd x Vd)] using the assumption of equal partition coefficients in the striatum and the cerebellum, and graphical analysis of the binding potential. Depending on the method of analysis, a 50-63% reduction of the binding to DA transporters in the caudate, and a 64-75% reduction in the putamen of the LND patients was observed compared to the normal control group. When LND patients were compared to Rett syndrome patients, similar reductions were found in the caudate (53-61%) and putamen (67-72%) in LND patients. Transporter binding in Rett syndrome patients was not significantly different from the normal controls. Finally, volumetric magnetic resonance imaging studies detected a 30% reduction in the caudate volume of LND patients. To ensure that a reduction in the caudate volume would not confound the results, a rigorous partial volume correction of the caudate time activity curve was performed. This correction resulted in an even greater decrease in the caudate-cerebellar ratio in LND patients when contrasted to controls. To our knowledge, these findings provide the first in vivo documentation of a dopaminergic reduction in LND and illustrate the role of positron emission tomography imaging in investigating neurodevelopmental disorders.

A strong association has been observed between [3H]zolpidem binding and the presence of gamma-aminobutyric acid (GABAA) receptor mRNA for alpha 1-, beta 2-, and gamma 2-subunits in specific brain regions. This correlates with observed sensitivity of individual neurons to zolpidem and ethanol in these same regions. Previous studies using homogenate binding approaches showed small alterations in [3H]zolpidem binding levels after chronic ethanol exposure. This study was undertaken to ascertain if there is regional specificity of the effects of chronic ethanol administration on [3H]zolpidem binding levels. Chronic ethanol administration induced small, but significant alterations in [3H]zolpidem (5 nM) binding in the inferior colliculus, substantia nigra, and the medial septum. [3H]Zolpidem binding was increased in the inferior colliculus and substantia nigra, and decreased in the medial septum. No significant differences in [3H]zolpidem binding were noted in any other brain area analyzed, including the cortex and cerebellum. These findings show that chronic ethanol administration has small effects on [3H]zolpidem binding, although they occur in a site-specific and bidirectional manner. Moreover, there is no correlation between changes in [3H]zolpidem binding and alterations in GABAA receptor subunit expression.

Previous investigations have suggested a relationship between zolpidem binding within specific brain regions and the ability of ethanol or zolpidem to enhance gamma-aminobutyric acid (GABA)-induced inhibition. The purpose of the present study was to extend our electrophysiological analysis to additional brain sites with high levels of zolpidem binding. In the brain regions chosen, red nucleus and globus pallidus, GABA-induced inhibition was shown to be enhanced by either ethanol or zolpidem on some, but not all, neurons. These findings led to the hypothesis that the effect of zolpidem on GABA-induced inhibition would predict the action of ethanol on responses to GABA for that neuron. When zolpidem and ethanol were applied individually to the same neurons in the red nucleus and globus pallidus, those neurons sensitive to zolpidem enhancement of GABA also were sensitive to ethanol. Conversely, if zolpidem did not enhance responses to GABA, ethanol did not enhance responses to GABA at these brain sites. A similar relationship between the abilities of zolpidem and ethanol to enhance GABA-induced inhibition was obtained in 90% of the neurons studied in the medial septum/diagonal band and ventral pallidum. These studies provide further support for the contention that the zolpidem-sensitive GABAA-benzodiazepine isoreceptor also responds to ethanol. Finally, the expression of GABAA subunit mRNAs was analyzed by polymerase chain reaction from micropunches of several brain regions that contain zolpidem binding sites and exhibit sensitivity to ethanol. Polymerase chain reaction analysis proved more sensitive than in situ hybridization in the detection of receptor subunit mRNAs. Several subunits (alpha 1, alpha 2, alpha 3, beta 2, beta 3 and gamma 2) were common to all brain regions in which ethanol and zolpidem enhanced GABA responses. GABAA receptor alpha 4/5, alpha 6, beta 1, gamma 1, gamma 3 and delta subunits were not consistently expressed in association with the presence of zolpidem binding. These data are consistent with the view that one native GABAA receptor to which zolpidem binds, and on which ethanol acts, contains the GABAA receptor subunits alpha 1, beta 2 and gamma 2; however, the present investigation did not preclude the possibility that other subunit combinations can contribute to ethanol and zolpidem enhancement of responses to GABA.

The prenatal and postnatal ontogeny of D1A and D2 dopamine receptors was assessed by in situ hybridization of messenger RNAs encoding the receptors and by radioligand binding autoradiography. On gestational day 14, signals for D1A and D2 dopamine receptor messages were observed in selected regions in ventricular and subventricular zones which contain dividing neuroblasts, and in intermediate zones that contain maturing and migrating neurons. Specifically, D1A and D2 dopamine receptor message was observed in the developing caudate-putamen, olfactory tubercle, and frontal, cingulate, parietal and insular cortices. Additionally, D1A dopamine receptor messenger RNA was found in the developing epithalamus, thalamus, hypothalamus, pons, spinal cord and neural retina; D2 dopamine receptor messenger RNA was also observed in the mesencephalic dopaminergic nuclear complex. Gene expression of D1A and D2 dopamine receptor subtypes in specific cells as they differentiate precedes dopamine innervation and implies that receptor expression is an intrinsic property of these neurons. The early expression of dopamine receptor messenger RNA suggests a regulatory role for these receptors in brain development. While the signal for both messages increased in the intermediate zones on gestational day 16, it decreased in the ventricular and subventricular zones, and was no longer apparent in these zones by gestational day 18. By gestational day 18, abundant D1A or D2 dopamine receptor messenger RNA was observed in cell groups similar in location to those observed in the adult brain. On gestational day 18, D1A dopamine receptor message was noted in the neural retina, anterior olfactory nucleus, the insular, prefrontal, frontal, cingulate, parietal and retrosplenial cortices, the olfactory tubercle, caudate-putamen, lateral habenula, dorsolateral geniculate nucleus, ventrolateral and mediolateral thalamic nuclei, and the suprachiasmatic and ventromedial nuclei of the hypothalamus. D2 dopamine receptor message was observed on gestational day 18 in the insular, prefrontal, frontal and cingulate cortices, the olfactory tubercle, caudate-putamen, ventral tegmental area, substantia nigra, and the intermediate lobe of the pituitary. At birth, expression of messenger RNA for both dopamine receptor subtypes in the striatum approximated that seen in mature rats. In contrast, D1A and D2 receptor binding, measured with [3H]SCH-23390 and [3H]raclopride, respectively, was low at birth and progressively increased to reach adult levels between days 14 and 21. The in situ hybridization data showing early prenatal expression of messenger RNA for the D1A and D2 dopamine receptors are consistent with the hypothesis that these receptors have a regulatory role in neuronal development.(ABSTRACT TRUNCATED AT 400 WORDS)

Alterations in brain activity patterns were assessed in response to swim stress by immunocytochemical detection of Fos-like immunoreactivity (Fos-LI) and high-resolution autoradiographic imaging of 14C-2-deoxyglucose (2-DG) uptake. The stress paradigm investigated was a classic behavioral screen for antidepressant drug activity, the forced swim test. One of the most pronounced effects produced by swim stress was an increase in 2-DG uptake and induction of Fos-LI in a restricted region of the lateral septal nucleus. Specific "limbic" cortical regions, including the medial prefrontal, ventrolateral orbital, and cingulate cortices, also exhibited both increased 2-DG uptake and expression of Fos-LI in response to swim stress. In the hypothalamic paraventricular nucleus of swim-stressed rats, Fos-LI was induced but no change in 2-DG uptake was apparent. Since the specific swim stress protocol used is a behavioral screen for antidepressant drugs, the effects of imipramine on stress-induced alterations in 2-DG uptake and induction of Fos-LI were examined. The stress-induced increase in 2-DG uptake in the lateral septum was blocked by treatment with imipramine, but treatment with imipramine had no effect on induction of Fos-LI in the same region. Neither 2-DG uptake nor Fos-LI expression was altered by imipramine treatment in the cortical regions influenced by swim stress. Administration of imipramine alone under basal conditions produced a robust induction of Fos-LI in the central nucleus of the amygdala and in the dorsal lateral subdivision of the bed nucleus of the stria terminalis. No effect of imipramine treatment on 2-DG uptake was apparent in these latter regions. The results provide insights into topographic patterns of brain activity associated with swim stress and neuroanatomically selective actions of imipramine. The different and complementary information obtained by assessment of Fos-LI and 2-DG uptake illustrates the utility of applying both functional mapping approaches to examine neuroanatomical correlates of behavioral states and drug treatment.

Behavioral activation following systemic administration of the N-methyl-D-aspartic acid receptor antagonist MK-801 [(+)-5-methyl-10,11-dihydroxy-5H-dibenzo(a,d) cyclohepten-5,10-imine; dizocilpine] was examined in unlesioned control and in neonatal-6-hydroxydopamine (OHDA) lesioned rats. Neonatal-6-OHDA lesioned animals were found more sensitive than control rats and female rats more sensitive than males to this MK-801-induced behavioral activation. CGS-19755, a competitive NMDA antagonist, also increased activity in neonatally lesioned animals. The increased activity level following MK-801 administration to neonatally lesioned rats was reduced, but not eliminated, by pretreatment with alpha-methyltyrosine, indicating that endogenous catecholamines were partially responsible for this action of MK-801. Furthermore, neither a D1- nor a D2-dopamine antagonist was totally effective alone in reducing MK-801-induced behavioral activation in the neonatally lesioned rats, but MK-801-induced activity was reduced to the level observed after alpha-methyltyrosine when both dopamine antagonists were administered in combination. In contrast to these results, alpha-methyltyrosine virtually eliminated the MK-801-induced activity in adult-lesioned rats. When individual behaviors induced by MK-801 were examined in neonatal-6-OHDA lesioned rats, MK-801 did not produce the same behaviors as L-dihydroxyphenylalanine or a D1- or D2-dopamine agonist. Whereas MK-801 had no major effect on most behaviors induced by specific D1- or D2-dopamine agonists, it blocked some behaviors produced after L-dihydroxyphenylalanine administration, including the self-injurious behavior. Repeated MK-801 treatment resulted in increasingly greater motor activity, but this was not related to increased D1-dopamine receptor sensitization. In support of a regional action of MK-801, MK-801 induced c-fos-like immunoreactivity in the cerebral cortex, but not in the nucleus accumbens or striatum. The action of MK-801 to increase c-fos-like immunoreactivity in cerebral cortex was reduced, but not blocked, by SCH-23390. Additionally, MK-801 reduced, but did not eliminate, D1-dopamine agonist induced c-fos-like immunoreactivity in striatum. These data suggest that MK-801 not only can facilitate dopamine release within specific brain regions, but has behavioral and functional actions distinct from dopamine agonists.

Our laboratory has previously shown that systemically administered ethanol inhibits NMDA-evoked electrophysiological activity in some, but not all, neurons in the medial septum. In the present report, it was found that ethanol, when applied locally via electro-osmosis, potently inhibited NMDA-evoked neuronal activity in a current-dependent manner in the inferior colliculus and hippocampus. In contrast, locally applied ethanol failed to inhibit NMDA-evoked activity in the lateral septum. The inhibition by ethanol of NMDA-evoked activity in the inferior colliculus was specific, in that ethanol failed to inhibit neuronal activity of the inferior colliculus evoked by the excitatory neurotransmitter glutamate. These findings indicate that ethanol can specifically inhibit NMDA-evoked activity in vivo via a local action, and that the ability of ethanol to inhibit NMDA-evoked activity varies regionally in brain. The possibility that these results are explained by the existence of two types of NMDA receptors, one sensitive to ethanol, the other insensitive to ethanol, is discussed.

The studies described will demonstrate that the subunit composition of a GABAA receptor allows ethanol to enhance responses to GABA. Since we have determined that ethanol will influence responses to glycine, nicotine and NMDA in some, but not all, neurons with receptors to these agonists, we hypothesize that specific receptor subtypes of these ligand-gated ion channels will be affected by ethanol.

The neuroanatomical distribution of binding sites for [3H]imipramine and [3H]citalopram was assessed by in vitro autoradiography in select regions of the rat and human forebrain. To determine involvement of serotonin-containing terminals in the binding of [3H]imipramine and [3H]citalopram, binding of these compounds was measured in rats after destroying serotonin-containing neurons with 5,7-dihydroxytryptamine (5,7-DHT). Treatment with this neurotoxin decreased serotonin content by 90% and reduced [3H]citalopram binding to a similar extent. These results demonstrate that [3H]citalopram binding is a reliable marker for serotonin-containing terminals. Binding of [3H]imipramine was reduced by only 15-35% after 5,7-DHT treatment. These latter results suggest that only a small fraction of [3H]imipramine binding to brain sections is associated with serotonergic terminals under standard conditions used in autoradiographic studies with the ligand. Dose-response effects of fluoxetine and desipramine on displacement of [3H]imipramine binding in forebrain regions indicate that the ligand labels predominantly high capacity, low affinity binding sites. To determine the utility of the rat brain as a model for [3H]imipramine and [3H]citalopram binding in the human brain, binding of the ligands was compared in human and rat hypothalamus, amygdala, and hippocampus. The pharmacological characteristics of [3H]imipramine and [3H]citalopram binding were similar in the rat and human brain. However, substantial species differences were observed in topographic patterns of [3H]imipramine binding within the hippocampus and hypothalamus. The distribution of [3H]citalopram binding sites within the amygdala and hypothalamus were also strikingly different in rats compared to humans. This work provides the first demonstration that marked species differences exist in the topography of serotonergic innervation and in the distribution of [3H]imipramine binding sites within the rat and human brain regions examined.

Effects of octanoic acid on monoamines and their acidic metabolites in the rat brain were analyzed by HPLC. Octanoic acid (1,000 mg/kg i.p.) elevated homovanillic acid levels by 54% in the caudate and 338% in the hypothalamus but increased 5-hydroxyindoleacetic acid (5-HIAA) levels in both the caudate and the hypothalamus by approximately 50% compared with the control. A lower dose of octanoic acid (500 mg/kg) increased 5-HIAA levels by 29% in the caudate and 20% in the hypothalamus. However, it did not produce any changes in the concentration of homovanillic acid in either the caudate or the hypothalamus. Treatment with octanoic acid also failed to change the level of dopamine, serotonin, and 3,4-dihydroxyphenylacetic acid in the caudate and the hypothalamus. The role of carrier-mediated transport in the clearance of 5-HIAA from the rabbit CSF was also evaluated in vivo by ventriculocisternal perfusion. Steady-state clearance of 5-HIAA from CSF exceeded that of inulin and was reduced in the presence of octanoic acid. Because this transport system in the choroid plexus is normally responsible for the excretion of the serotonin metabolite from the brain to the plasma, accumulation of endogenously produced organic acids in the brain, secondary to reduced clearance by the choroid plexus, could be a contributing factor in the development of encephalopathy in children with medium-chain acyl-CoA dehydrogenase deficiency who have elevated levels of octanoic acid systematically.

Previous work has demonstrated that the cortical sub-division of the inferior colliculus is capable of seizure genesis, so the present studies delineated the neuroanatomical extent of this seizure genesis using two mapping techniques, [14C]2-deoxyglucose (2-DG) uptake and electrical stimulation. When 4 inferior collicular seizures were elicited over a 5 min period, the stimulated side of the brain showed highly selective increases in the [14C]2-DG uptake in comparison to the unstimulated side. Although the substantial change in [14C]2-DG uptake occurred over the entire inferior collicular cortex, electrical stimulation mapping delineated a specific area capable of seizure genesis within the inferior collicular cortex. The electrical stimulation also identified a number of regions that would support electrically dependent seizure behaviors: the cuneiform nucleus, the ventrolateral inferior colliculus, portions of the dorsal central gray, and the peripeduncular nucleus. In concert, marked unilateral increases in [14C]2-DG uptake were found on the stimulated side in the peripeduncular/substantia nigra lateralis area, the medial geniculate and a specific region of the dorsal central gray. These studies verify the asymmetric nature of inferior collicular seizure genesis, identify areas of seizure modulation, and delineate a region in the inferior collicular cortex that modulates sensory-motor integration.

Expression of a D1 dopamine receptor was examined in the rat brain by using a combination of in situ hybridization and in vitro receptor autoradiography. Cells expressing D1 receptor mRNA were localized to many, but not all, brain regions receiving dopaminergic innervation. The highest levels of hybridization were detected in the caudate-putamen, nucleus accumbens, and olfactory tubercle. Cells expressing D1 receptor mRNA were also detected throughout the cerebral cortex, limbic system, hypothalamus, and thalamus. D1 receptor mRNA was differentially expressed in distinct regions of the hippocampal formation. Dentate granule cells were labeled in dorsal but not ventral regions, whereas the subicular complex was prominently labeled in ventral but not dorsal regions. Intermediate to high levels of D1 binding sites, but no hybridizing D1 receptor mRNA, were detected in the substantia nigra pars reticulata, globus pallidus, entopeduncular nucleus, and subthalamic nucleus. In these brain regions, which are involved in the efferent flow of information from the basal ganglia, D1 receptors may be localized on afferent nerve terminals originating in other brain regions. These results indicate that in addition to a role in control of motor function, the D1 receptor may also participate in the cognitive, affective, and neuroendocrine effects of dopaminergic neurotransmission.

Recent studies have demonstrated that ethanol blocks N-methyl-D-aspartate (NMDA) responses in vitro. In the present study, evidence is provided that ethanol, when administered by the systemic route to rats, also inhibits NMDA-evoked electrophysiological activity in vivo at behaviorally relevant doses. Ethanol, at doses in rats ranging from those producing minimal changes in spontaneous behavioral activity (0.75 g/kg) to those producing marked suppression of behavioral activity (2.5 g/kg), produced a dose-dependent inhibition of the ability of NMDA, when iontophoresed onto neurons of the medial septum (MS), to activate MS neurons. However, at all doses of ethanol tested, a proportion of MS neurons responded to ethanol with essentially complete inhibition of NMDA-evoked activity, whereas other MS neurons responded to ethanol with little or no inhibition of NMDA-evoked activity. By way of comparison, MK-801, a non-competitive NMDA antagonist, antagonized NMDA-evoked activity in all MS neurons tested. In contrast to the actions of ethanol, MK-801 increased, rather than decreased, behavioral activity even at doses that completely inhibited NMDA-evoked activity in all MS neurons tested. These latter findings provide evidence that inhibition of NMDA-evoked activity cannot account for all of the behavioral effects of ethanol. In conclusion, while the present results demonstrate for the first time that ethanol can inhibit NMDA-evoked neuronal activity in vivo, they also indicate that additional neural actions must contribute to ethanol's pharmacological profile.

Using high resolution autoradiographic techniques, the distribution of radioactivity in forebrain and brainstem was assessed after IV injection of 3H-impramine or 3H-desipramine. Results were compared with regional binding of the drugs to brain sections in vitro. Similar topographic binding of 3H-imipramine and 3H-desipramine was observed in vitro among brain regions, except in the paraventricular nucleus of the hypothalamus and locus coeruleus, where binding was greater for 3H-desipramine. For both 3H-desipramine and 3H-imipramine, some brain regions that exhibited high binding in vitro also showed high accumulation after in vivo injection. However, certain regions that contained high densities of binding sites for the antidepressant drugs as measured by in vitro binding showed very low accumulation of radioactivity after in vivo treatment. Such regions included the dentate gyrus of the hippocampus, layer 1 of piriform cortex, caudate-putamen, pontine and midbrain central gray, and cerebellar granular layer. Compared to in vitro binding of the drugs, the distribution of imipramine and desipramine in vivo appears more anatomically selective. For imipramine, primary sites of action in vivo, as indicated by the topographic distribution in brain, appear to be the locus coeruleus, hippocampus, lateral septal nucleus, and amygdala. For desipramine, the greatest accumulation in vivo was found in the locus coeruleus, paraventricular nucleus of the hypothalamus, and anterior thalamic nuclei.

Because of uncertainty concerning the interaction of ethanol with gamma-aminobutyric acid (GABA) receptor-mediated events, the present work was designed to investigate the effect of ethanol on GABA transmission in the rat septal area using behavioral and electrophysiological techniques. Microinjection of the GABAA agonist muscimol into the medial septal area (MSA) enhanced, and bicuculline administration antagonized, ethanol-induced impairment of the aerial righting reflex. Microinjection of these drugs into the lateral septum (LSi) did not influence this measure of ethanol-induced sedation. Furthermore, intraseptal injections of muscimol or bicuculline in saline-treated rats had no effect on the aerial righting reflex. These data suggest that the MSA plays a critical modulatory role in the sedative actions of ethanol. To assess the effect of ethanol on muscimol responses in the MSA and LSi at the cellular level, GABA was applied by iontophoresis to rhythmically bursting neurons of the MSA and to cells in the LSi. The magnitude of the resultant inhibition by GABA on these cells was assessed before and after systemic administration of ethanol. Ethanol enhanced GABA-mediated inhibition of MSA neural activity, but did not alter GABA-mediated inhibition of cellular activity in the LSi. In contrast, the inhibition of cellular activity in the MSA, caused by a maximally effective concentration of the benzodiazepine flurazepam, was not altered by ethanol. Other work in the MSA demonstrated that electrical stimulation of the fimbria caused an inhibition of ongoing single unit activity that was reduced by concurrent application of bicuculline. The duration of this electrically elicited inhibition in the MSA was enhanced after ethanol injection and then recovered to base-line levels. In addition, ethanol (1.5 mg/kg) caused an enhancement of the inhibition induced by nipecotic acid, a GABA uptake inhibitor. These findings demonstrate that GABA-mediated neural inhibition is enhanced by ethanol in the MSA but not the LSi, indicating that the actions of ethanol on GABA-induced inhibition can be site specific. It is proposed that the cellular action of ethanol may depend upon a specific molecular composition of the GABA receptor complex which may vary at selected sites in the brain.

Dopamine (DA) and cholecystokinin octapeptide carboxy-terminal (CCK-8) have been found to coexist in some mesolimbic neurons. The present investigation was undertaken in order to study the biochemical and behavioral interactions between CCK-8 and some central monoaminergic pathways. The action of the sulfated form of CCK-8 (10 micrograms/10 microliter intracerebroventricularly) on DA turnover in nucleus accumbens, olfactory tubercles and corpus striatum of the rat was determined after DA synthesis inhibition with alpha-methyl-p-tyrosine (250 mg/kg i.p.). Also, CCK-8 action (1-30 micrograms intracisternally) on DA synthesis was assessed by measuring accumulation of dihydroxyphenylalanine (DOPA) after DOPA-decarboxylase inhibition with NSD-1015 (m-hydroxybenzylhydrazine, 100 mg/kg i.p.). The contents of DA and its main metabolites, dihydroxyphenylacetic acid and homovanillic acid, together with serotonin and its main metabolite, 5-hydroxyindoleacetic acid (5-HIAA), were measured in different brain areas after direct injection of CCK-8 into the ventral tegmental area (A10) or nucleus accumbens. Further, the effect of CCK-8 on amphetamine-induced locomotion and apomorphine-induced stereotypies was studied along with changes in spontaneous locomotion and rearing after CCK-8 injection into the ventral tegmental area and nucleus accumbens. No consistent statistically significant effects of CCK-8 on biochemical or behavioral assessments on measures of DA function were observed. However, injection of high doses of CCK-8 into the ventral tegmental area significantly decreased levels of 5-HIAA in the nucleus accumbens, olfactory tubercles and striatum.

Neurotensin (NT), an endogenous tridecapeptide, is heterogeneously distributed in the central nervous system. The present study examined the effects of physiologically and behaviorally active doses of NT (1--100 micrograms intracisternally) on dopamine, serotonin and their primary metabolites as well as accumulation of dopa after inhibition of dopa decarboxylase. NT was shown to increase dopa accumulation when compared with saline treatment, suggesting that dopamine synthesis was increased. In accord with this view, NT also caused a dose-dependent increase in homovanillic acid and dihydroxyphenylacetic acid, the major metabolites of dopamine, in several brain areas (striatum, olfactory tubercles, nucleus accumbens, frontal cortex and hypothalamus). Interestingly, the increase in homovanillic acid was greater than that for dihydroxyphenylacetic acid. In striatum, an initial increase in dopamine content after 30 micrograms of NT was followed by an increase and a subsequent decrease of dopamine metabolites. Several other neuropeptides (Met-enkephalin, cholecystokinin-8, thyrotropin releasing hormone, substance P and d-Arg9-NT), at doses equimolar to 30 micrograms of NT, did not affect dopamine metabolites, whereas certain others (beta-endorphin and bombesin) increased their concentration in some brain areas. Except for the highest dose of NT, measures of serotonergic function were not affected by NT or any of the other neuropeptides.

Previous work by the authors has established decreased brain ATP concentration after a combined hypoxic-hypotensive episode. This study was undertaken to determine what changes, if any, occur in brain norepinephrine (NE), dopamine (DA), and serotonin (5HT) concentration in association with hypoxia and hypotension, and to correlate any observed changes with simultaneously measured ATP concentrations. Rats were subjected to a 30-minute period of hypoxia (F102 = 0.075) and hemorrhagic hypotension (MAP = 30 mm Hg) and then resuscitated. Significant increases (P less than .05) in cortical 5HT were observed at a time when ATP concentration was significantly (P less than .005) reduced. Additional experiments were conducted on rats depleted of 5HT by prior treatment with 5,7-dihydroxytryptamine. Equal decreases in ATP concentration were measured, and the cardiovascular response to hypoxic hypotension in 5HT-depleted rats was similar to that in 5HT-intact rats. We conclude that the increased 5HT after hypoxic hypotension does not cause the decreased ATP concentration, nor does 5HT play a major role in cardiovascular homeostasis under the conditions of this experiment.

Locomotor activity induced by d-amphetamine was found to be potentiated by food deprivation, a tryptophan-free diet, p-chlorophenylalanine and drugs proposed to antagonize serotonin receptors in brain. Administration of L-tryptophan 1 hour prior to d-amphetamine injection was found to antagonize the enhanced response to d-amphetamine in starved rats and in rats which had tryptophan removed from their diet. However, tryptophan did not block the potentiated response to d-amphetamine in animals pretreated with p-chlorophenylalanine. These findings suggested that the antagonism of d-amphetamine-induced activity by tryptophan in starved rats and rats fed a tryptophan-free diet was not due to a nonspecific depressant effect of the amino acid. Since accumulation of d-amphetamine and its metabolites was not affected by any of the treatments which enhanced its activity, it seems unlikely that an alteration in the metabolism of d-amphetamine can explain these findings. The present work provides additional support for the view that serotonergic fibers play an important role in the actions of d-amphetamine.

Bombesin is a tetradecapeptide heterogenously distributed in the mammalian brain. Bombesin (45 micrograms) given intracisternally (IC) to unanesthetized rats increased the accumulation of dihydroxyphenylalanine (DOPA) in striatum, olfactory tubercles and hypothalamus after DOPA-decarboxylase inhibition, thus indicating an increased dopamine synthesis. A dose-dependent increase in dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), the principal dopamine metabolites, was seen in several brain areas 1 hr after IC injection of bombesin (0-60 micrograms). In striatum and olfactory tubercles HVA increased more than DOPAC with a maximal increase after 30-45 micrograms. In a time-course experiment a biphasic change of dopamine metabolites was observed in the olfactory tubercles with an actual decrease in metabolite levels 4 hr after 60 micrograms IC bombesin injection. Co-administration of bombesin and naloxone (8 mg/kg IP) or ethanol (2.25 g/kg IP) did not affect the increase in dopamine metabolites seen after bombesin alone. The action of IC administered bombesin on dopamine function was most pronounced in hypothalamus indicating a neuroendocrine regulatory of the peptide.

Evidence has been reviewed which strongly suggests that TRH functions as a neurotransmitter or neuromodulator in the mammalian central nervous system. Both the peptide and its receptor sites are located in the brain. Furthermore, it has protein actions to modify the effects of many neuropharmacological agents and can itself cause alterations in functions mediated by the CNS. Data clearly indicate that many of these actions. of TRH are not dependent on the pituitary- thyroid axis. Various studies of neurotransmitter interactions with TRH have provided evidence that noradrenergic, serotonergic, GABAergic, and cholinergic systems may be influenced by TRH or mediate some of its actions. More than likely, other transmitters will be implicated as the complex actions of TRH are more thoroughly investigated and understood. One set of experiments suggested that TRH may be localized serotonergic fibers. Such findings provide strong support for the view that TRH has a role in the physiology of the CNS. In spite of the progress that has been made, there are several questions to be answered about mechanisms of synthesis, storage, release, and inactivation of TRH. Furthermore, physiological studies would be greatly facilitated if a specific antagonist of TRH actions were available. Controversy still exists about active forms of TRH in situ and the methods by which TRH can be measured in tissue. Future investigations which resolve these difficulties and questions should facilitate our understanding of the role of TRH in brain function and its complex interactions with other neural mechanisms.

Biochemical studies have shown that the ability of erythrosine to inhibit dopamine uptake into brain synaptosomal preparations is dependent on the concentration of tissue present in the assay mixture. Thus, the finding that erythrosine inhibits dopamine uptake (which, if true, would provide a plausible explanation of the Feingold hypothesis of childhood hyperactivity) may simply be an artifact that results from nonspecific interactions with brain membranes. In addition, although erythrosine given parenterally (50 milligrams per kilogram) did not alter locomotor activity of control of 6-hydroxydopamine-treated rats, erythrosine (50 to 300 milligrams per kilogram) attenuated the effect of punishment in a "conflict" paradigm.

The selectivity and specificity of fenmetozole (DH-524) [2(3,4-dichlorophenoxy-methyl)2-imidazole HCl] as an antagonist of the actions of ethanol were examined. Fenmetozole (15--30 g/kg) reduced ethanol-induced impairment of the aerial righting reflex without changing blood or brain ethanol content, indicating that the antagonistic actions of fenmetozole were not de to change in the pharmacokinetics of ethanol. Since fenmetozole also reduced aerial righting reflex impairment due to phenobarbital, chlordiazepoxide, and halothane, this action of fenmetozole was not specific to ethanol. In mice, both the ethanol-induced increase in locomotor activity at 2.0 g/kg and the decrease caused by 4.0 g/kg were antagonized by fenmetozole. In addition, fenmetozole attenuated the ethanol-induced reduction in cerebellar cyclic guanosine monophosphate (cGMP) content, but the drug also significantly elevated cGMP levels in this tissue when given alone. Fenmetozole did not alter ethanol-induced increases in punished drinking in a conflict test, except at a high dose which alone decreased both punished and unpunished responding. Fenmetozole also failed to precipitate ethanol withdrawal-like reactions when given to physically-dependent, intoxicated rats. Thus, the antagonistic action of fenmetozole against ethanol would not seem to be related to a specific receptor interaction but rather may be the result of a physiological antagonism.

Thyrotropin-releasing hormone (TRH) causes a significant increase in rat cerebellar guanosine 3',5'-monophosphate (cGMP) content after parenteral administration. A smaller but still significant increase in cGMP was also observed in brain stem, whereas no significant changes were observed in cGMP in other gross brain regions or in adenosine 3',5'-monophosphate in any brain region. TRH also caused a similar increase in cerebellar cGMP content in hypophysectomized rats indicating that the increase is independent of an intact pituitary. The time course of cGMP elevation in the cerebellum after administration of 10 mg/kg of TRH i.v. showed a peak at 2.5 to 5.0 min followed by a less rapid decrease. The time course of TRH immunoreactivity in the same cerebellar homogenates roughly paralleled these changes. Only those TRH analogs which were previously shown to antagonize pentobarbital sleeping time in mice caused an elevation in cGMP content in the cerebellum. TRH also caused a significant increase in cerebellar cGMP in rats pretreated with phenobarbital and chlordiazepoxide. The TRH-induced increase in cerebellar cGMP was not affected by cerebellar climbing fiber lesions caused by 3-acetylpyridine nor blocked by haloperidol, suggesting that TRH acts by mechanisms different from harmaline or apomorphine in raising cerebellar cGMP.

Antagonism of ethanol-induced depression of locomotion was observed after intracisternal injection of thyrotropin releasing hormone (TRH) and bicuculline methiodide (BICM), as well as after microinjection of these drugs into the medial septum. The present investigation compared the behavioral and physiological consequence of administering TRH and BICM into the medial septum, inferior colliculus and substantia nigra to quantitate the similarities between these compounds. BICM produced a major increase in locomotor activity when injected into the medial septum and stereotypies when injected into the substantia nigra, suggesting that GABA-containing neurons have widespread influences on motor function. The wild running and seizure activity observed after BICM injection into the inferior colliculus was also consistent with this latter view. The marked increase in rectal temperature observed when BICM was injected into the medial septum may also implicate GABAergic mechanisms in temperature control at this brain site. TRH produced no such behavioral or physiological changes when administered into these three sites. Thus, this work strongly suggests that TRH does not exert a widespread action as a GABA antagonist because TRH did not produce the same changes induced by BICM. The actions of BICM and TRH to antagonize ethanol-induced depression when microinjected into the medial septum suggests that this brain area may be a critical site for the depressant action of ethanol.

Resting with CO2 stimulated respiration were measured by means of a whole body plethysmograph in rats lightly anesthetized with halothane. The respiratory effects of different doses of the serotonin precursor 5-HTP, and the serotonin agonist 5-methoxy-N,N-dimethyltryptamine were studied as well as the effects of a serotonin antagonist methysergide and p-chlorophenylalanine, an inhibitor of serotonin synthesis. The serotonergic agonists decreased tidal volume and minute volume in a dose dependent manner and produced a respiratory acidosis. The respiratory depressant effect was antagonized by methysergide, and the serotonergic antagonist and synthesis inhibitor alone stimulated respiration. Rats given intraventricular 5-methoxy-N,N-dimethyltryptamine also evidenced a decrease in tidal volume, and this response was greater in animals given 5,7-dihydroxytryptamine. It seems likely that CNS serotonin receptors are involved in the control of both basal and CO2 stimulated respiration.

Aminophylline-induced stimulation of respiration in halothane-anesthetized rats was abolished in rats given 6-hydroxydopamine and desmethylimipramine neonatally to selectively destroy central nervous system dopamine nerve terminals. The respiratory stimulation was also prevented by prior administration of dopamine receptor antagonists haloperidol and cisflupenthixol. The combination of apomorphine and aminophylline produced an additive stimulatory effect on respiration to values greater than those observed with either drug alone. Animals which received pargyline and 5,7-dihydroxytryptamine neonatally to selectively destroy central nervous system serotonin nerve terminals appeared to be more sensitive to the stimulatory effects of aminophylline on respiration when tested as adults. The depressant effect of 5-methoxy-N, N-dimethyl-tryptamine on respiration was slightly enhanced in the presence of aminophylline. These data suggest that the respiratory response to aminophylline is the summation of the effects of this drug on central nervous system dopamine on serotonin neuronal systems.

In this study the effect of administering 6-hydroxydopamine (6-OHDA) intracisternally on brain catecholamine content and fluorescence patterns of cerebellar processes was examined. It was found that intracisternal injection of 6-OHDA resulted in widely diverging effects depending upon the dose of 6-OHDA, age of the animal upon injection and the length of the post injection interval. Small doses of 6-OHDA (3 and 10 microgram) selectively depleted telencephalic and upper brain stem NE while larger doses of 6-OHDA (30 and 100 microgram) infringed on dopaminergic as well as noradrenergic neurons. In addition, the lower doses of 6-OHDA, but not the higher ones, led to an approximately two-fold accumulation of NE in the lower brain stem and cerebellum. Morphological observations suggested that the cerebellar norepinephrine accumulation after 10 microgram 6-OHDA was attributable primarily to an invasion of noradrenergic processes into the cerebellum.

Following codepletion of endogenous serotonin (5-HT, greater than 90%) and thyrotropin-releasing hormone (TRH, 66%) by neonatal treatment with the serotonergic neurotoxin, 5,7-dihydroxytryptamine (DHT), a 33% (n = 12, P less than 0.01) increase in specific TRH receptor binding was observed in adult rat spinal cord (SC) homogenates. A 20-21% increase in TRH receptors was also observed in the medulla/pons (MP) (n = 12, P less than 0.05) and midbrain (MB) (n = 12, P less than 0.02), but no changes were detected in 6 rostral brain regions. The depletion of 5-HT after DHT-treatment was also accompanied by a 34-42% increase in 5-HT1 binding in the SC, MP and MB. Eadie-Hofstee analysis revealed that the changes in TRH receptor levels observed after DHT-lesions were due to an increase in receptor number rather than any significant changes in receptor affinity. Chronic treatment of adult rats with the 5-HT-depleting drugs, p-chlorophenylalanine (PCPA) and reserpine, produced a 90-97% decrease in 5-HT in the SC, MP and MB and elevated 5-HT1 binding above controls in these tissues. However, neither drug treatment caused any significant alterations in the levels of TRH or its receptors in any of these tissues. In conclusion, these results have provided further support for the coexistence of 5-HT and TRH in the MP and SC and revealed possible new areas of such colocalization in the MB. Furthermore, these data have demonstrated that only DHT-treatment, as apposed to PCPA or reserpine, can produce long-lasting codepletion of 5-HT and TRH with simultaneous compensatory up-regulation of their receptor systems in the SC and other caudal tissues.

Intracisternal NGF or anti-NGF has been found to produce no long-term major alterations in central norepinephrine (NE) or dopamine levels when administered to neonatal rats. While NGF and anti-NGF were found to produce significant changes in brain NE content within one week of treatment, changes in central NE were no longer detectable at 30 days of age. Modification of the growth response of the central adrenergic neurons following 6-OHDA treatment was also not affected by NGF or anti-NGF when evaluated 3 weeks after treatment. However, centrally administered anti-NGF did induce a loss of peripheral NE terminals, which was attributed to leakage of the anti-NGF from the central injection site.

Serotonergic neurons throughout the brain were destroyed by early postnatal treatment of rats with an intracisternal injection of 5,7-dihydroxytryptamine (5,7-DHT), as demonstrated with biochemical measurements of serotonin and immunocytochemical localization of serotonin-containing neurons. Using these methods, it was shown that approximately 75-98% of serotonergic neurons underwent cell death in rats which were treated on day 3. In contrast, intracisternal administration of 5,7-DHT in adult rats led to the loss of distal serotonergic terminals without apparent loss of the cell bodies. Desipramine pretreatment prevented significant effects of 5,7-DHT on noradrenergic neurons.

Hemophilus influenzae is the most common cause of bacterial meningitis in children, and a high percentage of survivors are at risk for long-term sequelae. To explore the mechanisms responsible for these sequelae, a neonatal rat model was used to define the behavioral, electrophysiological, and biochemical changes following meningitis. Three days after inoculation of 6-day-old rats with a minimum of 1 X 10(7) colony-forming units of a virulent Hemophilus influenzae, type b, cerebrospinal fluid and blood were cultured to confirm the presence of meningitis and bacteremia, respectively. At this time, forebrain norepinephrine and dopamine levels were significantly elevated in meningitic rats when standardized on a wet-weight basis. No changes in brain serotonin or heart norepinephrine levels could be found in the 9-day-old rats. No residual changes were found in steady-state concentrations of norepinephrine or dopamine in surviving adult rats. However, survivors that had had meningitis as neonates showed significant impairment in active and passive avoidance learning tasks and demonstrated a significantly higher level of activity during a habituation period in circular photocell activity cages. No change in the flinch-jump threshold was detected. Brainstem auditory evoked potentials showed delays of various waves in 3 of 10 Hemophilus influenzae type b-treated adult rats tested. These rats also exhibited markedly augmented locomotory responses to d-amphetamine (1 mg/kg), suggesting a long-lasting perturbation of central monoamine neuronal transmission.

The diverse behavioral and biochemical effects induced by ethanol suggest that ethanol exerts differential effects on the CNS. When the neuroactive amino acids, glycine, glutamate, aspartate, GABA and taurine, were measured in the cortex, striatum, hippocampus, midbrain, and brain stem of acute or chronic ethanol-treated rats, site specific changes were observed for glutamate, glycine, and aspartate. No changes were found for GABA or taurine. Upon in vivo application, it was found that the microinjection of thyrotropin-releasing hormone (TRH, 500 ng) into the medial septum significantly shortened ethanol's impairment of the righting reflex, while microinjection of muscimol (30 ng) markedly potentiated ethanol's impairment of the righting reflex. When these studies are combined with previous work showing that microinjection of muscimol (30 ng) into the inferior colliculus blocks audiogenically induced seizures in ethanol-withdrawn rats, the convergence implies that specific sites in the CNS may modulate certain actions of ethanol. Therefore, we propose that the medial septum and inferior colliculus can be used as in vivo models to study the acute and chronic actions of ethanol, respectively.

In the present investigation, the neuropharmacology of 1,3 butanediol (1,3-BD) was compared with that of ethanol. Acute i.p. administration of equimolar doses of 1,3-BD or ethanol to rats impaired the aerial righting reflex, attenuated the suppressive effect of punishment on drinking behavior, lowered blood pressure, caused a concomitant reduction in the content of guanosine 3',5'-monophosphate in the cerebellum and reduced ethanol withdrawal reactions. Although these data suggested that ethanol and 2,3-BD were of similar potency, the brain content of 1,3-BD was only 33% of that of ethanol after treatment with equimolar doses, suggesting a greater central nervous system (CNS) potency for 1,3-BD. In rats treated chronically with ethanol to produce physical dependence, 1,3-BD was more potent than ethanol in inhibiting the hyperexcitability observed upon ethanol withdrawal. Furthermore, chronic administration and withdrawal of 1,3-BD caused CNS hyperexcitability in rats that was characteristic of physical dependence. Despite these similarities, there were clear differences in the actions of ethanol and 1,3-BD. In mice, locomotor stimulation caused by ethanol was not observed after 1,3-BD. Furthermore, while 1,3-BD did not alter the concentration of luteinizing hormone in plasma, equivalent doses of ethanol markedly reduced the concentration of this hormone. These data indicate that like ethanol, 1,3-BD depresses CNS activity and induces physical dependence, but has less effect on plasma luteinizing hormone concentration than ethanol.

The significance of central noradrenergic, dopaminergic and serotonergic neural systems for the locomotor stimulant effects of methylphenidate was investigated in the rat. In order to study the role of brain catecholamines, rats were pretreated with reserpine (2.5 mg/kg) followed 24 hrs later by treatment with alpha-methyltyrosine (25 mg/kg) or U-14,624 (75 mg/kg), a dopamine-beta-hydroxylase inhibitor. In these experiments, methylphenidate stimulated motor activity was antagonized by alpha-methyltyrosine and enhanced after treatment with U-14,624, suggesting that release of newly synthesized dopamine is important to a locomotor stimulant action of methylphenidate. Evidence implicating brain serotonin in the actions of methylphenidate was obtained in rats pretreated with pargyline or p-chlorophenylalanine (PCPA). Administration of pargyline 1 hr prior to methylphenidate was found to reduce the locomotor activity induced by methylphenidate and this was antagonized by pretreatment with low doses of PCPA. Higher doses of PCPA caused a significant elevation of methylphenidate induced activity which could be reduced by 5-hydroxytryptophan. Destruction of serotonergic neurons with 5,7-dihydroxytryptamine also potentiated methylphenidate induced locomotion. These latter findings suggest that serotonergic fibers have an inhibitory function in brain. These results are discussed in relation to the possible mechanism by which methylphenidate may act in hyperkinesis.

Substance P, thyrotropin-releasing hormone (TRH) and serotonin are putative neurotransmitters which have been proposed to co-exist in some brain neurons. Our previous immunocytochemical and biochemical studies have demonstrated that 85-100% of all serotonin neurons are destroyed following neonatal 5,7-dihydroxtryptamine (5,7-DHT) treatment. In this study, we have determined the effect of neonatal 5,7-DHT and desipramine (DMI) treatment on the biochemical content and immunocytochemical localization of substance P and TRH throughout the brain. Interestingly, we have observed that virtually all substance P- and TRH-immunoreactive cells in the ventral pons-medulla are destroyed by the neurotoxin. However, peptide-containing neurons in other regions were not affected. Additionally, we measured the peptide content and found that TRH is significantly decreased in the spinal cord (-50%) and pons-medulla (-20%), but not in other brain regions. Substance P content was not significantly altered in any region, even after a greater than 90% reduction of serotonin. These data indicate that the co-localized substance P and TRH forms a small proportion of the total peptide in brain.

The latency, duration, hindlimb kick frequency, and total activity components of the post-decapitation reflex (PDR) were measured in the rat using a movement-sensitive transducer. Reduction of brain and spinal cord norepinephrine (NE) caused by neonatal administration of 6-hydroxydopamine (6-OHDA) or 5,7-dihydroxytryptamine, which also reduced brain serotonin, decreased all components of the PDR. Depletion of serotonin or dopamine alone reduced the vigor of the reflex, suggesting that these pathways can influence the PDR but are not essential for the response. Lesions of neurons in the Locus coeruleus, made electrolytically or with 6-OHDA, decreased the intensity of the PDR, with the 6-OHDA-induced lesion being more effective. Depletion of forebrain NE terminals with 6-OHDA did not alter the PDR, consistent with a critical involvement of spinal noradrenergic fibers. The PDR was also decreased by phentolamine and prazosin, but not by propanolol, suggesting an involvement of alpha-adrenergic receptors in the response. This hypothesis was further supported by the finding that the efficacy of a variety of drugs (such as tricyclic antidepressants, phenothiazines, and anti-hypertensive compounds) for blocking the reflex was apparently related to their affinity for alpha-adrenergic receptors. Thus, the PDR is dependent on noradrenergic fibers in the spinal cord and may provide a simple screen for drugs with suspected alpha-adrenergic blocking properties or for agents that disrupt the function of central noradrenergic fibers.

When rats were pretreated with ethanol (3.0 g/kg, IP), subsequent microinjection of thyrotropin-releasing hormone (TRH) (500 or 1000 ng) into the medial septum, 30 minutes later, significantly shortened the time necessary for the rats to regain their righting reflex. Conversely, microinjection of TRH into the nucleus accumbens (1000 ng/side) or the area of the raphe obscurus (1000 ng) had no effect on ethanol-induced depression, although both of these structures mediate specific TRH effects in the CNS. In order to determine if this antagonism was due to a specific TRH interaction, TRH Fab fragments were microinjected into the medial septum just prior to the microinjection of TRH. Under these conditions, TRH did not alter ethanol's depressant actions. Finally, this TRH antagonism of ethanol-induced depression appears attributable to a net increase in neuronal activity, because electrical stimulation (160 microA, 120 Hz, 1.5 msec duration) of the medial septum antagonized ethanol's impairment of the righting reflex. These results are discussed in relationship to a potential CNS site for the action of ethanol.

After injection of 2-deoxyglucose (2-DG) labeled with tritium or carbon-14, autoradiograms were produced by thaw-mounting 4 micron frozen sections of rat brains on nuclear emulsion-coated slides. The results show that the distribution of radioactivity among different brain regions was similar and that the resolution at the regional topographical level was virtually identical for both compounds. The resolution obtained with the thaw-mounting of thin frozen sections onto nuclear emulsion was considerably greater than the resolution demonstrated in published results in the literature, when carbon-14 or tritium-labeled 2-deoxyglucose were used with 20 micron frozen sections and X-ray film or tritium-sensitive film. The results indicate that section thickness, detection medium and intimacy of contact between section and photographic emulsion influence resolution. At the regional level, the detection medium apparently influences resolution to a greater extent than the energy differences of the beta particles emitted from 14C or 3H. Although diffusion of radiolabeled 2-deoxyglucose and metabolites during the thaw-mounting process precludes single cell resolution of these autoradiograms, the improvement of visualizing regional topographic detail demonstrates that the described technique is a valuable approach with which to study regional 2-DG uptake.