Biomedical subjects

D A Morilak

Publications and source records attributed to D A Morilak.

At least 19 recordsLinked to original sources

PubMed2005

Noradrenergic modulation of cognitive function in rat medial prefrontal cortex as measured by attentional set shifting capability.

The brain noradrenergic system is thought to facilitate neuronal processes that promote behavioral activation, alertness, and attention. One region in which norepinephrine may exert such effects is the medial prefrontal cortex, which has been implicated in many cognitive functions including arousal, attention, motivation, working memory, response inhibition, and behavioral flexibility. The present study addressed the modulatory influence of noradrenergic neurotransmission in medial prefrontal cortex on cognitive function in rats, as measured by performance in an attentional set shifting task. In experiment 1, we tested effects of increasing and decreasing brain noradrenergic neurotransmission by systemic administration of the alpha2-adrenergic autoreceptor antagonist and agonist drugs, atipamezole and clonidine, respectively. Atipamezole pretreatment significantly improved performance on the stages of the attentional task requiring an extradimensional shift in attention, and those involving stimulus reversals, whereas clonidine had no effect at any stage. In experiment 2, we then tested effects of microinjecting alpha1- or beta-adrenergic receptor antagonists into medial prefrontal cortex on the enhancement of performance on the extradimensional task produced by atipamezole. The atipamezole-induced enhancement of performance on the extradimensional set shifting task was blocked by alpha1-, but not beta-adrenergic receptor antagonists in medial prefrontal cortex. Neither antagonist alone had any effect on extradimensional set shift performance in the absence of atipamezole-induced enhancement. These results indicate that elevating noradrenergic activity at alpha1-receptors in medial prefrontal cortex facilitates cognitive performance of rats in an attentional set-shifting task, which may contribute to the role of norepinephrine in behavioral state changes such as arousal, or to the beneficial cognitive effects of psychotherapeutic drugs that target noradrenergic neurotransmission.

Adrenergic alpha-Agonists↗

PubMed2005

Norepinephrine release in medial amygdala facilitates activation of the hypothalamic-pituitary-adrenal axis in response to acute immobilisation stress.

Activation of the brain noradrenergic system during stress plays an important integrative function in coping and stress adaptation by facilitating transmission in many brain regions involved in regulating behavioural and physiological components of the stress response. The medial amygdala (MeA) has been implicated in modulation of stress-induced activation of the hypothalamic-pituitary-adrenal (HPA) axis, and MeA is a target of innervation from brainstem noradrenergic neurones. However, it is not known whether, and to what extent, activation of the ascending noradrenergic innervation of MeA might modulate stress-induced adrenocorticotropic hormone (ACTH) secretion. In the first experiment in this study, we measured extracellular norepinephrine (NE) levels in MeA using in vivo microdialysis. The concentration of NE in dialysate samples collected in MeA was elevated by more than three-fold over baseline in response to acute immobilisation stress, providing evidence of a possible modulatory role for NE in the MeA during stress. This potential role was then assessed in the second experiment by measuring changes in the elevation of plasma ACTH concentration induced by acute immobilisation stress immediately following bilateral microinjections of alpha1- or beta-adrenergic receptor antagonists directly into MeA. Compared to vehicle-injected controls, the alpha1-receptor antagonist benoxathian dose-dependently and significantly attenuated the ACTH response to acute stress, whereas combined beta1/beta2-receptor blockade in MeA had only a modest effect. These results indicate that MeA does play a role in the stress response, and support the hypothesis that stress-induced activation of NE release in MeA, acting primarily through alpha1 receptors, facilitates activation of the HPA axis in response to acute stress.

Acute Disease↗

PubMed2005

Chronic intermittent cold stress sensitises the hypothalamic-pituitary-adrenal response to a novel acute stress by enhancing noradrenergic influence in the rat paraventricular nucleus.

Chronic intermittent cold stress sensitises activation of the hypothalamic-pituitary-adrenal (HPA) axis by novel acute stress. We have shown that enhanced noradrenergic function in limbic forebrain contributes to HPA sensitisation. In the present study, we investigated whether chronic intermittent cold also induced changes in noradrenergic function in the paraventricular nucleus (PVN), the primary mediator of the HPA stress response. Rats were exposed to chronic intermittent cold (7 days, 6 h per day, 4 degrees C). On the day after final cold exposure, there were no differences in baseline plasma ACTH, but the peak ACTH response to 30 min of acute immobilisation stress was greater in cold-stressed rats compared to controls. Bilateral microinjection of the alpha(1)-adrenergic receptor antagonist benoxathian into the PVN reduced acute stress-induced adrenocorticotrophic hormone (ACTH) levels by approximately 25% in controls. Furthermore, in cold-stressed rats, all of the sensitisation of the ACTH response was blocked by benoxathian, to a level comparable to benoxathian-treated controls. In a second study using microdialysis to measure norepinephrine release in the PVN, there were no differences in either baseline or acute stress-induced increases in norepinephrine release in the PVN of cold-stressed rats compared to controls. Thus, in a third study, we tested potential alterations in postsynaptic alpha(1)-receptor sensitivity after chronic cold stress. Dose-dependent activation of ACTH secretion by microinjection of the alpha(1)-adrenergic receptor agonist, phenylephrine, into the PVN was significantly enhanced in cold-stressed rats compared to controls. Thus, the sensitised HPA response to acute stress after chronic intermittent cold exposure is at least partly attributable to an enhanced response to alpha1-adrenergic receptor activation in the PVN. Chronic stress-induced plasticity in the acute stress response may be important for stress adaptation, but may also contribute to pathophysiological conditions associated with stress. Thus, understanding the neural mechanisms underlying such adaptations may help us understand the aetiology of such disorders, and contribute to the future development of more effective treatment or prevention strategies.

Adrenergic alpha-Agonists↗

PubMed2004

Induction of FOS expression by acute immobilization stress is reduced in locus coeruleus and medial amygdala of Wistar-Kyoto rats compared to Sprague-Dawley rats.

Activation of the brain noradrenergic system during acute stress is thought to play an important integrative function in coping and stress adaptation by facilitating transmission in many brain regions involved in regulating behavioral and physiologic components of the stress response. Compared with outbred control Sprague-Dawley (SD) rats, inbred Wistar-Kyoto (WKY) rats exhibit an exaggerated hypothalamic-pituitary-adrenal (HPA) response as well as increased susceptibility to certain forms of stress-related pathology. However, we have also shown previously that WKY rats exhibit reduced anxiety-like behavioral reactivity to acute stress, associated with reduced activation of the brain noradrenergic system. Thus, to understand better the possible neurobiological mechanisms underlying dysregulation of the stress response in WKY rats, we investigated potential strain differences in stress-induced neuronal activation in brain regions that are both involved in regulating behavioral and neuroendocrine stress responses, and are related to the noradrenergic system, either as targets of noradrenergic modulation or as sources of afferent innervation of noradrenergic neurons. This was accomplished by visualizing stress-induced expression of Fos immunoreactivity in the paraventricular nucleus of the hypothalamus, lateral bed nucleus of the stria terminalis, central nucleus of the amygdala, and medial nucleus of the amygdala (MeA), as well as the noradrenergic nucleus locus coeruleus (LC). Stress-induced Fos expression was found to be decreased in the LC and MeA of WKY rats compared with similarly stressed SD rats, whereas no strain differences were observed in any of the other brain regions. This suggests that strain-related differences in activation of the MeA may be involved in the abnormal neuroendocrine and behavioral stress responses exhibited by WKY rats. Moreover, as the MeA is both an afferent as well as an efferent target of the brainstem noradrenergic system, reduced MeA activation may either be a source of reduced noradrenergic reactivity seen in WKY rats, or possibly a consequence. Nonetheless, understanding the mechanisms underlying altered stress reactivity in models such as the WKY rat may contribute to a better understanding of stress-related psychopathologies such as depression, post-traumatic stress disorder or other anxiety disorders.

Amygdala↗

PubMed2002

Modulatory effects of norepinephrine in the lateral bed nucleus of the stria terminalis on behavioral and neuroendocrine responses to acute stress.

The brain noradrenergic system is activated by stress, and modulates the activity of forebrain regions involved in behavioral and neuroendocrine responses to stress, such as the lateral bed nucleus of the stria terminalis (BSTL). This region of the limbic forebrain receives dense noradrenergic innervation, and has been implicated in both anxiety and regulation of the hypothalamic-pituitary-adrenal axis. We hypothesized that stress-induced release of norepinephrine in the BSTL modulates anxiety-like behavioral responses to stress and activation of the hypothalamic-pituitary-adrenal stress axis. Using microdialysis, we showed that release of norepinephrine was increased in the BSTL of male Sprague-Dawley rats during immobilization stress. In the next experiment, we then microinjected noradrenergic antagonists into the BSTL immediately prior to acute immobilization stress to examine noradrenergic modulation of behavioral stress reactivity. Either the alpha(1)-receptor antagonist benoxathian, or a cocktail of beta(1)- and beta(2)-receptor antagonists (betaxolol+ICI 118,551) blocked the anxiety-like reduction in open-arm exploration on the elevated plus-maze, but not the reduction in social behavior induced in the social interaction test. In a third experiment, benoxathian reduced plasma levels of adrenocorticotropic hormone following stress, but beta-receptor antagonists had no effect. From these results we suggest that stress-induced norepinephrine release acts on both alpha(1)- and beta-receptors in the BSTL to facilitate anxiety-like behavioral responses on the plus-maze but not the social interaction test, and modulates hypothalamic-pituitary-adrenal axis activation via alpha(1)-receptors only. Together with previous results in which adrenergic antagonists in central amygdala attenuated behavioral responses on the social interaction test but not the plus-maze, these observations suggest the two behavioral tests measure different dimensions of stress reactivity, and that norepinephrine facilitates different components of the stress response by region- and receptor-specific mechanisms.

Acute Disease↗

PubMed2002

Stress reactivity of the brain noradrenergic system in three rat strains differing in their neuroendocrine and behavioral responses to stress: implications for susceptibility to stress-related neuropsychiatric disorders.

The brain noradrenergic system is activated by stress, modulating the activity of forebrain regions involved in behavioral and neuroendocrine responses to stress. In this study, we characterized brain noradrenergic reactivity to acute immobilization stress in three rat strains that differ in their neuroendocrine stress response: the inbred Lewis (Lew) and Wistar-Kyoto (WKY) rats, and outbred Sprague-Dawley (SD) rats. Noradrenergic reactivity was assessed by measuring tyrosine hydroxylase mRNA expression in locus coeruleus, and norepinephrine release in the lateral bed nucleus of the stria terminalis. Behavioral measures of arousal and acute stress responsivity included locomotion in a novel environment, fear-potentiated startle, and stress-induced reductions in social interaction and open-arm exploration on the elevated-plus maze. Neuroendocrine responses were assessed by plasma adrenocorticotropic hormone. Compared to SD, adrenocorticotropic hormone responses of Lew rats were blunted, whereas those of WKY were enhanced. The behavioral effects of stress were similar in Lew and SD rats, despite baseline differences. Lew had similar elevations of tyrosine hydroxylase mRNA, and initially greater norepinephrine release in the lateral bed nucleus of the stria terminalis during stress, although both noradrenergic responses returned toward baseline more rapidly than in SD rats. WKY rats showed depressed baseline startle and lower baseline exploratory and social behavior than SD. However, unlike the Lew or SD rats, WKY exhibited a lack both of fear potentiation of the startle response and of stress-induced reductions in exploratory and social behavior, indicating attenuated stress responsivity. Acute noradrenergic reactivity to stress, measured by either tyrosine hydroxylase mRNA levels or norepinephrine release, was also attenuated in WKY rats. Thus, reduced arousal and behavioral responsivity in WKY rats may be related to deficient brain noradrenergic reactivity. This deficit may alter their ability to cope with stress, resulting in the exaggerated neuroendocrine responses and increased susceptibility to stress-related pathology exhibited by this strain.

Animals↗

PubMed2000

Effects of acute restraint stress on tyrosine hydroxylase mRNA expression in locus coeruleus of Wistar and Wistar-Kyoto rats.

Norepinephrine (NE) is thought to play a role in the stress response, and may be involved in stress-related psychopathological conditions such as depression or anxiety. Heterogeneity in individual responses to the same stressor suggest that a genetic susceptibility to the effects of stress may contribute to such pathology. To address possible mechanisms underlying this genetic aspect of the stress response, we examined acute stress-induced changes in mRNA expression for several components of the NE system in the locus coeruleus (LC) and adrenal medullae of stress-susceptible Wistar-Kyoto (WKY) rats and their parent Wistar (W) strain. Expression of tyrosine hydroxylase (TH), NE transporter (NET) and alpha(2A) receptor mRNA were measured in the LC by in situ hybridization 30 min and 2 h after the onset of 30 min restraint stress. Adrenal TH mRNA was measured by slot blots. No basal differences were observed for any measure, but in the LC, expression of TH mRNA increased by 40% in W rats at 30 min (n=8, p<0.05) and returned toward baseline by 2 h, while WKY rats showed only a non-significant 29% increase at 2 h. In contrast, adrenal TH mRNA expression increased in WKY rats at 2 h (n=3, p<0.05), with no significant change in W rats. NET and alpha(2A) mRNA were unaltered by restraint stress in both strains. Differences in the stress-reactivity of TH gene expression in the central and peripheral noradrenergic systems may be related to differences in behavioral coping strategies and autonomic responsivity to stress in these strains, and suggest that differences in noradrenergic reactivity may contribute to genetic susceptibility to stress-related pathology.

Animals↗

PubMed2000

Changes in tyrosine hydroxylase mRNA expression in the rat locus coeruleus following acute or chronic treatment with valproic acid.

Valproate has proven effective in treating bipolar disorder. Though some biochemical effects of valproate are rapid, mood-stabilizing effects can take weeks, suggesting that regulatory changes in gene expression in brain neurotransmitter systems may be involved. Given a presumed role for norepinephrine (NE) in bipolar disorder, as well as the actions of mood-stabilizing drugs, we examined changes in mRNA expression for tyrosine hydroxylase (TH), the NE transporter (NET) and alpha 2A autoreceptor in the rat locus coeruleus after valproate treatment. TH mRNA increased slightly (16%) following acute treatment, and more so after chronic valproate treatment (26%), while neither NET nor alpha 2A mRNA expression changed. Further, chronic valproate treatment attenuated the elevation in TH mRNA expression induced in the LC in response to acute restraint stress. Both acute and chronic valproate treatment attenuated restraint stress-induced elevations in plasma ACTH secretion. These observations suggest that the therapeutic effects of valproate may involve regulatory alterations in TH message expression in the brain, and attenuation of stress-reactivity of the central noradrenergic system and the hypothalamic-pituitary-adrenal axis.

Adrenocorticotropic Hormone↗

PubMed1999

Effects of chronic antidepressant treatments on serotonin transporter function, density, and mRNA level.

To investigate functional changes in the brain serotonin transporter (SERT) after chronic antidepressant treatment, several techniques were used to assess SERT activity, density, or its mRNA content. Rats were treated by osmotic minipump for 21 d with the selective serotonin reuptake inhibitors (SSRIs) paroxetine or sertraline, the selective norepinephrine reuptake inhibitor desipramine (DMI), or the monoamine oxidase inhibitor phenelzine. High-speed in vivo electrochemical recordings were used to assess the ability of the SSRI fluvoxamine to modulate the clearance of locally applied serotonin in the CA3 region of hippocampus in drug- or vehicle-treated rats. Fluvoxamine decreased the clearance of serotonin in rats treated with vehicle, DMI, or phenelzine but had no effect on the clearance of serotonin in SSRI-treated rats. SERT density in the CA3 region of the hippocampus of the same rats, assessed by quantitative autoradiography with tritiated cyanoimipramine ([(3)H]CN-IMI), was decreased by 80-90% in SSRI-treated rats but not in those treated with phenelzine or DMI. The serotonin content of the hippocampus was unaffected by paroxetine or sertraline treatment, ruling out neurotoxicity as a possible explanation for the SSRI-induced decrease in SERT binding and alteration in 5-HT clearance. Levels of mRNA for the SERT in the raphe nucleus were also unaltered by chronic paroxetine treatment. Based on these results, it appears that the SERT is downregulated by chronic administration of SSRIs but not other types of antidepressants; furthermore, the downregulation is not caused by decreases in SERT gene expression.

Adrenergic Uptake Inhibitors↗

PubMed1999

Expression of alpha1D adrenergic receptor messenger RNA in oxytocin- and corticotropin-releasing hormone-synthesizing neurons in the rat paraventricular nucleus.

The paraventricular nucleus of the hypothalamus contains a number of intermingled populations of neuroendocrine cell groups involved in the hormonal stress response, including cells synthesizing corticotropin-releasing hormone and oxytocin. Ascending noradrenergic afferents to the paraventricular nucleus, acting through alpha1 adrenergic receptors, are thought to play a role in stress-induced activation of the hypothalamic-pituitary-adrenal axis. We have previously demonstrated that, of the three known alpha1 adrenergic receptor subtypes, messenger RNA for the alpha1D subtype is the most prominently expressed in the paraventricular nucleus. Thus, regulation of the expression of this receptor may be important in modulation of the stress response. It is currently unknown, however, which populations of stress-related neuroendocrine cells in the paraventricular nucleus express alpha1 receptors, or whether the excitatory influence of norepinephrine in stress is exerted directly on neurons expressing oxytocin or corticotropin-releasing hormone. Thus, in the present study, we used dual in situ hybridization, combining a digoxigenin-labeled riboprobe encoding the rat alpha1D adrenergic receptor with radiolabeled riboprobes for oxytocin or corticotropin-releasing hormone, to determine the degree to which these neurons in the paraventricular nucleus express alpha1D adrenergic receptors. In sections through the rostral and mid-level paraventricular nucleus, nearly all (>95%) oxytocin neurons also expressed alpha1D messenger RNA. In contrast, the populations of corticotropin-releasing hormone- and alpha1D-expressing cells overlapped only partially, with most alpha1D expression situated more laterally. A subset (37%) of the neurons expressing corticotropin-releasing hormone also expressed alpha1D messenger RNA, and these were found almost entirely within the region of overlap in the lateral aspect of the medial parvocellular region. These observations support a direct role for alpha1 receptors in regulation of oxytocin secretion. Expression of alpha1D messenger RNA in distinct subsets of cells synthesizing corticotropin-releasing hormone may also help to clarify contradictory and inconsistent observations in the literature regarding the role of norepinephrine in the stress response, and may account for a presumed stressor-specific role for norepinephrine in activation of the hypothalamic-pituitary-adrenal axis.

Animals↗

PubMed1998

Reduced noradrenergic tone to the hypothalamic paraventricular nucleus contributes to the stress hyporesponsiveness of lactation.

Lactation in mammals is accompanied by a marked decrease in stress responsiveness that we previously attributed, in part, to a reduction in noradrenergic (NA) innervation of hypothalamic paraventricular nucleus (PVN) neurons controlling neuroendocrine stress responses. In the present study, we compared in-vivo PVN catecholamine secretion by microdialysis between nonlactating and lactating females and tested the effects of NA alpha-1 and alpha-2 receptor antagonists (corynanthine and idazoxan, respectively) on the acute stress response of lactating and virgin female rats. To determine if PVN alpha-adrenoreceptor density, affinity, or synthesis, changes as a function of lactation, we performed receptor autoradiography, Scatchard analysis and in situ hybridization of alpha-adrenoreceptors. Densitometric analysis of the alpha-adrenoreceptors in the supraoptic nucleus (SON) was used to evaluate changes in magnocellular neurons. Endogenous PVN norepinephrine release under basal conditions was lower in lactating females than in females who had their pups removed for 2 days, and microdialysate concentrations of adrenaline and MHPG were attenuated in lactating females. Alpha-2 adrenoreceptor density in the PVN showed a significant decrease from lactation day 3 to lactation days 10-12 and a reduction to 40% of virgin controls on days 10-20 of lactation. A similar pattern was observed for the SON. The affinity of hypothalamic alpha-2 adrenoreceptors was reduced as a function of lactation. Alpha-1 adrenoreceptor density in the PVN and in the hypothalamus rose as a function of lactation, although the affinity of these receptors was not altered. In contrast, alpha-1D adrenoreceptor subtype mRNA expression in the PVN decreased in middle lactating females (day 10) compared to virgins. Intracerebroventricular (i.c.v.) application of idazoxan, significantly increased the ACTH response to swim stress in virgin females, but had the opposite effect in lactating females. In contrast, i.c.v. corynanthine treatment significantly decreased the ACTH response in virgins, but not in lactating females. Overall, these data suggest that the secretion of NA in the PVN is reduced during lactation, and that the ability of PVN parvocellular neurons to respond to changes in synaptic NA levels (i.e. after stress) is also altered.

Adrenergic alpha-Antagonists↗

PubMed1997

Distribution of alpha1A adrenergic receptor mRNA in the rat brain visualized by in situ hybridization.

Norepinephrine has been implicated in a number of physiological, behavioral, and cellular modulatory processes in the brain, and many of these modulatory effects are attributable to alpha1 adrenergic receptors. At least three alpha1 receptor subtypes have been identified by molecular criteria, designated alpha1A, alpha1B, and beta1D. The distributions of alpha1B and alpha1D receptor mRNA expression in rat brain have been described previously, but the cDNA for the rat alpha1A receptor has only recently been cloned and characterized. In the present study, we used a radiolabelled riboprobe derived from the rat alpha1A receptor cDNA to describe the distribution of alpha1A message expression in the rat brain. The highest levels of alpha1A adrenergic receptor mRNA expression were seen in the olfactory bulb, tenia tectae, horizontal diagonal band/magnocellular preoptic area, zona incerta, ventromedial hypothalamus, lateral mammillary nuclei, ventral dentate gyrus, piriform cortex, medial and cortical amygdala, magnocellular red nuclei, pontine nuclei, superior and lateral vestibular nuclei, brainstem reticular nuclei, and several cranial nerve motor nuclei. Dual in situ hybridization combining a radioactive riboprobe for choline acetyltransferase mRNA with a digoxigenin-labeled alpha1A riboprobe in the fifth and seventh cranial nerve motor nuclei showed that the alpha1A mRNA is expressed in cholinergic motor neurons. Prominent alpha1A hybridization signal was also seen in the neocortex, claustrum, lateral amygdala, ventral cochlear nucleus, raphe magnus, and in the ventral horn of thoracic spinal cord. This overall pattern of expression, considered in comparison with that previously described for the other alpha1 adrenergic receptor subtypes, may shed light on the different roles of the alpha1 receptors in mediating the neuromodulatory effects of norepinephrine in processes such as arousal, neuroendocrine control, sensorimotor regulation, and the stress response.

Animals↗

PubMed1997

alpha1B adrenoceptors in rat paraventricular nucleus overlap with, but do not mediate, the induction of c-Fos expression by osmotic or restraint stress.

A role has been suggested for hypothalamic alpha1 adrenoceptors in the acute stress-induced activation of the hypothalamic-pituitary-adrenal axis. Using a polyclonal antiserum against the rat alpha1B adrenergic receptor protein, we have demonstrated alpha1B receptor immunoreactivity in neurons and especially in punctate cell processes in the rat paraventricular nucleus. The distribution of alpha1B receptor immunoreactivity overlapped in part with the distributions of c-Fos immunoreactivity induced in the paraventricular nucleus by either restraint stress or hypertonic saline administration. However, intraperitoneal pretreatment with the alpha1 receptor antagonist prazosin (0.5 or 5.0 mg/kg) failed to attenuate stress-induced c-Fos expression in the paraventricular nucleus. Prazosin also failed to attenuate the secretion of corticosterone following restraint stress. Thus, we conclude that neither acute secretory activity nor activation of gene transcriptional responses mediated by c-Fos in the hypothalamic pituitary adrenal axis following these stressors are dependent upon hypothalamic alpha1 adrenergic receptors.

Adrenergic alpha-1 Receptor Antagonists↗

PubMed1997

Co-localization of alpha1D adrenergic receptor mRNA with mineralocorticoid and glucocorticoid receptor mRNA in rat hippocampus.

Many adaptive changes occur in response to chronic or repeated stress, involving complex regulatory interactions between central stress-related afferents and the central components of the hypothalamo-pituitary-adrenal (HPA) axis. One change associated with chronic stress is an attenuation of corticosteroid receptor-mediated feedback inhibition of the HPA axis, a process thought to involve corticosteroid receptors in the hippocampus. A prominent stress-related central afferent that innervates the hippocampus and that may participate in the regulation of the HPA axis is the central noradrenergic system. Previous evidence suggests that alpha1 adrenergic receptors may down-regulate hippocampal corticosteroid receptors, and may thus contribute to stress-induced facilitation of HPA responses. In the present study, we used combined nonisotopic and radioisotopic in situ hybridization to examine the overlapping expression and co-localization of mRNA encoding the post-synaptic alpha1D adrenergic receptor subtype, the major alpha1 subtype expressed in hippocampus, with mRNA for the two corticosteroid receptor subtypes, type I (mineralocorticoid receptor, MR) and type II (glucocorticoid receptor, GR) in rat hippocampal neurons. We observed overlapping distributions and an extensive degree of co-localization of alpha1D receptor mRNA with both corticosteroid receptor subtype messages, establishing an anatomical substrate by which these two receptor systems may directly regulate each other. The potential interaction between co-localized adrenergic and corticosteroid receptors in hippocampus may contribute to stress-induced alterations in the HPA response to subsequent stress.

Animals↗

PubMed1994

Neurons expressing 5-HT2 receptors in the rat brain: neurochemical identification of cell types by immunocytochemistry.

The serotonin2 (5-HT2) receptor has been implicated in a number of behavioral and physiological processes. It may also play a role in cellular development and differentiation, and represents a site of action of hallucinogens and certain psychotherapeutic drugs. To better understand the functions and regulation of the 5-HT2 receptor, we have undertaken a series of studies in which we attempted to identify the specific cell types that express the receptor. This was accomplished using a variety of double-labeling strategies with an antibody we raised against the rat 5-HT2 receptor protein. In this review, we recount of some of our previously published findings and present some new data in which we identify subpopulations of cholinergic neurons in the brainstem and gamma-aminobutynic acid (GABA)ergic interneurons in the cortex that express 5-HT2 receptor immunoreactivity. Developmentally, the appearance of 5-HT2 receptor immunoreactivity occurs relatively late in teh ontogeny of the cells in which it is expressed, mostly in the early postnatal period. This argues against a significant role for this receptor in early development, though it may participate in some aspect of terminal differentiation. We discuss the significance of the cell-type-specific and temporal expression of the 5-HT2 receptor in the context of current hypotheses of neuropsychiatric disorders such as schizophrenia.

Animals↗

PubMed1993

5-HT2 receptor immunoreactivity on cholinergic neurons of the pontomesencephalic tegmentum shown by double immunofluorescence.

The serotonin-2 (5-HT2) receptor subtype is implicated in several behavioral and physiological processes, and may be the site of action of hallucinogens and certain psychotherapeutic drugs. To better understand the function and regulation of 5-HT2 receptors, it is necessary to determine the specific brain regions and cell types expressing them. By double immunofluorescence using a polyclonal antibody raised against the rat 5-HT2 receptor in conjunction with an antibody against choline acetyltransferase (ChAT), the synthetic enzyme for acetylcholine, we have shown that cholinergic neurons in the rat laterodorsal and pedunculopontine tegmental nuclei express 5-HT2 receptors. In contrast, there was little co-localization of 5-HT2 and ChAT immunoreactivity in neurons of the basal forebrain or striatum, even though the 5-HT2- and ChAT-positive cells in these regions overlapped extensively. These findings are discussed in relation to the potential interaction between cholinergic and serotonergic systems in sleep regulation, hallucinogenesis and the pathophysiology of neuropsychiatric disorders.

Acetylcholine↗

PubMed1993

Production and characterization of a specific 5-HT2 receptor antibody.

A synthetic peptide was used to generate antibodies against the rat serotonin-2 (5-HT2) receptor. The peptide corresponds to a unique sequence from the N-terminal extracellular portion of the receptor protein (antibody = Ab 5HT2-N). This peptide was chosen based on its theoretical antigenic index and for specificity to the 5-HT2 receptor. In dot blot analysis, antisera detected 2 ng-2 micrograms of synthetic peptide at dilutions of 1/200-1/20,000. COS-7 cells transiently transfected with a eukaryotic expression vector containing the 5-HT2 cDNA displayed intense immunoreactivity with crude and affinity-purified Ab 5HT2-N. In contrast, no immunoreactivity was seen in control experiments when: (1) non-transfected or vector transfected COS-7 cells were used; (2) pre-immune sera was substituted for primary antisera; (3) primary antisera was omitted; or (4) antiserum was pre-adsorbed to 10 microM synthetic peptide. Immunohistochemical analysis of sections of perfused rat brain revealed intense immunolabelling of a subset of neurons in regions of the ventral forebrain, dorsal hippocampus, striatum, cerebral cortex, and laterodorsal tegmental nucleus (LDT). An especially dense band of small cells was seen in layer 2 of pyriform cortex. There was a very high concentration of labelled cells in the laterodorsal tegmental nucleus. In situ hybridization histochemistry with a 5-HT2 antisense cRNA riboprobe showed a pattern of hybridization in forebrain similar to the pattern of immunolabelling with Ab 5HT2-N. Western blot analysis of proteins extracted from the LDT revealed a single protein species reacting with the antibody. This reactivity is not present in the pre-immune sera and is blocked by the synthetic antigen.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1993

Immunocytochemical localization and description of neurons expressing serotonin2 receptors in the rat brain.

Serotonin2 receptors have been implicated in a variety of behavioral and physiological processes, as well as a number of neuropsychiatric disorders. To specify the brain regions and specific cell types possessing serotonin2 receptors, we conducted an immunocytochemical study of the rat brain using a polyclonal serotonin2 receptor antibody. Perfusion-fixed rat brain sections were processed for immunocytochemistry and reactivity was visualized using an immunoperoxidase reaction. Numerous small, round neurons were heavily labeled in the granular and periglomerular regions of the olfactory bulb. Heavy labeling of medium-sized multipolar and bipolar neurons was also seen in olfactory regions of the ventral forebrain, including the anterior olfactory nucleus and olfactory tubercle. Other regions of the basal forebrain exhibiting high levels of immunoreactivity were the nucleus accumbens, ventral pallidum, Islands of Calleja, fundus striatum and endopyriform nucleus. Immunoreactive neurons were also seen in the lateral amygdala. A dense band of small, round cells was stained in layer 2 of pyriform cortex. In neocortex, a very sparse and even distribution of bipolar and multipolar neurons was seen throughout layers II-VI. A much more faintly labeled population of oval cells was observed in the deep layer of retrosplenial and posterior cingulate cortex, and in the granular layer of somatosensory frontoparietal cortex. A moderate number of medium bipolar and multipolar cells were scattered throughout the neostriatum, and a moderate number of pyramidal and pyramidal-like cells were seen in the CA fields of the hippocampus. Diencephalic areas showing immunolabeling included the medial habenula and anterior pretectal nucleus, with less labeling in the ventral lateral geniculate. In the hindbrain, two dense populations of large multipolar cells were heavily labeled in the pedunculopontine and laterodorsal tegmental nuclei, with lesser labeling in the periaqueductal gray, superior colliculus, spinal trigeminal nucleus and nucleus of the solitary tract. Based on the distribution, localization and morphology of immunoreactive neurons in these regions, we hypothesize that subpopulations of serotonin2 containing cells may be GABAergic interneurons or cholinergic neurons. Further, the observed distribution suggests that the physiological effects of serotonin acting through serotonin2 receptors are mediated by a relatively small number of cells in the brain. These observations may have strong functional implications for the pharmacological treatment of certain neuropsychiatric disorders.

Animals↗