Photoperiod regulation of mineralocorticoid receptor mRNA expression in hamster hippocampus

Hippocampal mineralocorticoid receptor mRNA expression was increased in male hamsters exposed to 18 days of short photoperiod relative to animals maintained under long day illumination (p<0.05). Short day hamsters were also characterized by increased weight gain, and heavier adrenal glands (p<0.05). The larger adrenals showed selective increases in the widths of the zonae reticularis and glomerulosa (p<0.001). Incidences of torpor and reduced body temperature were observed in the short day animals. No changes were found in reproductive organ weights, systolic blood pressure, open-field behavior, or stress levels of plasma corticosteroids. We conclude that the hamster brain–adrenal axis responds rapidly to changes in photoperiod, raising the possibility that this axis is a primary mediator of shortened photoperiod responses.     

Corticosterone alters N-methyl-D-aspartate receptor subunit mRNA expression before puberty

Stress and stress hormones alter the expression of mRNA for the NR1, NR2A and NR2B subunits of the N-methyl-D-aspartate (NMDA) receptor in brain regions associated with the stress response. Early life stress contributes to the risk and pathophysiology of mental illness. Examining how stress hormones modulate NMDA receptor subunit gene expression before and after pubertal onset will further contribute to the understanding of how stress during adolescence relates to adult mental illness. Using in situ hybridization histochemistry, we measured NR1, NR2A and NR2B mRNA expression in the hippocampus and in the hypothalamic paraventricular nucleus (PVN) of rats that had undergone adrenalectomy (ADX) or sham surgery before or after puberty. Some ADX rats received corticosterone pellets that released either normal or stress levels of corticosterone for 14 days prior to sacrifice. There was a significant increase in NR1 subunit mRNA expression throughout the subfields of the hippocampus and in the PVN of ADX prepubertal rats. However, similar changes in hippocampal NR1 expression were not observed in postpubertal ADX rats. Pre- and postpubertal ADX rats implanted with a high-dose corticosterone pellet had decreased expression of PVN NR1 mRNA. Only prepubertal rats had an increase in dentate gyrus NR2A mRNA and CA3 region NR2B mRNA following high-dose replacement. These results provide evidence that glucocorticoids have differential effects on the regional expression of mRNA NMDA receptor subunits and elucidate a window during adolescence in which the NR1, NR2A and NR2B genes are responsive to glucocorticoids.     

Circadian variations in expression of the trkB receptor in adult rat hippocampus

The expression of brain-derived neurotrophic factor (BDNF) in the central nervous system (CNS) and the expression of its high-affinity trkB receptor on neuron surfaces are known to depend on neuron activity. The expression of BDNF (mRNA and protein) and trkB mRNA shows circadian oscillations in rat hippocampal homogenates. We investigated circadian variations in trkB expression in specific areas of the adult rat hippocampal formation by immunohistochemistry. In sets of two experiments performed in the spring, 39 2-month-old male Wistar rats were accustomed to a 12-h light-12-h dark cycle for 2 weeks. Three animals were then sacrificed every 4 h. Forty-micrometer-thick coronal sections of hippocampal formation were obtained and processed for trkB immunohistochemistry. Cell staining intensity was assessed by image analysis of different hippocampal areas on five sections per animal. Circadian rhythmicity was evaluated by the cosinor method. Statistically significant circadian variations in trkB expression were found in dentate gyrus, entorhinal cortex, and the CA3 and hilar regions of the hippocampus, with highest expression during the first half of the dark (activity) period. These findings suggest a relationship between trkB expression and the physiological neuronal activation of wakefulness. TrkB receptor expression in the hippocampal regions studied was continuous and changes were gradual over the 24-h cycle, suggesting that more complex regulatory mechanisms also intervened.     

Glucocorticoid and mineralocorticoid receptor mRNA expression in squirrel monkey brain

Corticosteroids have been implicated in hippocampal atrophy in patients with severe psychiatric disorders, but little is known about receptor expression for corticosteroids in human or nonhuman primate brain. Both the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) were surveyed in this study of squirrel monkey brain using in situ hybridization histochemistry. Regions of high GR mRNA levels included CA1 and CA2 of hippocampus, dentate gyrus, paraventricular hypothalamus, lateral geniculate, lateral>medial amygdala, and cerebellum. Western analysis confirmed that GR immunoreactivity in squirrel monkey brain tissue most likely reflects the alpha isoform. Regions of high MR mRNA levels included all hippocampal pyramidal cell fields, dentate gyrus granule cell layer, lateral septum, medial>lateral amygdala, and to a lesser extent, cerebellum. Low levels of MR were also expressed in caudate and putamen. Receptor expression for corticosteroids in deep brain structures and the hippocampal formation was similar to that previously reported in rodents, but GR and MR mRNA were expressed at higher levels in squirrel monkey cerebral cortex. GR expression was evident in all cortical layers, particularly the pyramidal cell-rich layers II/III and V. MR expression was restricted to the more superficial cortical layers, and was only moderately represented in layer V. Laminar patterns were apparent in all regions of cortex for GR expression in squirrel monkeys, but low MR mRNA levels were found in dorsomedial prefrontal cortex (PFC). Different subregional distributions and distinctive laminar patterns suggest specialized functions or coordinated interactions between GR and MR mediated functions in primate PFC.     

Lesion of the substantia nigra pars compacta downregulates striatal glutamate receptor subunit mRNA expression

This is a study of the effect of the unilateral administration of dopamine (DA) in the pars compacta of the substantia nigra (SN) of the rat on striatal glutamate receptor subunit (GluR1, GluR2 and NMDAR1) gene expression determined by in situ hybridization. The location of the nigral lesion was determined by tyrosine hydroxylase (TH) immunohistochemistry and its extent by the striatal DA and 3,4-dihydroxyphenylacetic acid (DOPAC) concentrations. The DA-induced lesions produce significant bilateral reductions in the expression of GluR1 and NMDAR1 subunit mRNA in the medio-lateral striatum, whereas the expression of striatal GluR2 receptors was not changed. The reduction in GluR1 and NMDAR1 subunit mRNA may be the consequence of glutamatergic hyperactivity developed in the presence of a damaged nigro-striatal system and these may be associated with the genesis of some neurodegenerative diseases.     

Glucocorticoid and mineralocorticoid receptor expression in the human hippocampus in major depressive disorder

Approximately 50% of mood disorder patients exhibit hypercortisolism. Cortisol normally exerts its functions in the CNS via binding to mineralocorticoid receptors (MR) and glucocorticoid receptors (GR). Both MR and GR are highly expressed in human hippocampus and several studies have suggested that alterations in the levels of MR or GR within this region may contribute to the dysregulation in major depressive disorder (MDD). Studies have also shown functional heterogeneity across the hippocampus, with posterior hippocampus preferentially involved in cognitive processes and anterior hippocampus involved in stress, emotion and affect. We therefore hypothesize that GR and MR expression in hippocampus of control and MDD patients may vary not only with disease, but also with regional specificity along the anterior/posterior axis. Student's t-test analysis showed decreased expression of MR in the MDD group compared to controls in the anterior, but not the posterior hippocampus, with no significant changes in GR. Linear regression analysis showed a marked difference in MR:GR correlation between suicide and non-suicide patients in the posterior hippocampus. Our findings are consistent with previous reports of hippocampal corticosteroid receptor dysregulation in mood disorders, but extend those findings by analysis across the anterior/posterior axis of the hippocampus. A decrease in MR in the anterior but not posterior hippocampus of MDD patients emphasizes the important functional role of the anterior hippocampus in neuroendocrine regulation in humans.     

Corticosterone regulates expression of BDNF and trkB but not NT-3 and trkC mRNA in the rat hippocampus

Corticosterone has profound effects on growth, differentiation, and synaptic transmission of hippocampal neurons by activation of mineralocorticoid receptors (MRs) and glucocorticoid receptors (GRs). In the present study we tested if neurotrophins can be implicated in these effects. For this purpose we injected 30, 300, and 1,000 μg corticosterone s.c. (per kg body weight) in adrenalectomized rats and measured the mRNA levels of brain-derived neurotrophic factor (BDNF), tyrosine receptor kinase (trk)B, neurotrophin (NT)-3, and trkC in hippocampal cell fields at 6 hr after steroid administration by in situ hybridization. NT-3 and trkC mRNA did not show significant changes in any hippocampal region after the various doses of conticosterone. BDNF mRNA decreased after corticosterone administration dose dependently, resulting in a maximal suppression of 35, 20, and 50% in dentate gyrus, CA3, CA1, respectively. Interestingly, trkB responded to corticosterone in an inverted U-shaped fashion in CA3 and dentate gyrus: the low dose of corticosterone increased trkB mRNA expression in both regions by approximately 30%, while the effect of the two higher doses was not different from the vehicle injected controls. In conclusion, we found differential effects of low and high doses of corticosterone on BDNF and trkB expression in hippocampus, which suggests involvement of a coordinated MR- and GR-mediated action. J. Neurosci. Res. 48:334–341, 1997. © 1997 Wiley-Liss, Inc.     

Early auditory deprivation alters expression of NMDA receptor subunit NR1 mRNA in the rat auditory cortex

The expression of NMDA receptor NR1 subunit mRNA was studied in rat auditory cortex (AC) on different postnatal days using digoxigenin-labeled oligonucleotide probes. The results showed that NR1 expression increased from birth to postnatal day 35 (P35) and remained constant until P56. The most significant increases occurred between P7 and P14. Changes in NR1 mRNA expression in rats subjected to monaural hearing deprivation on P7, P21, P35, and P49 were examined on P56. Between P7 and P21, when the rat auditory system was still in a critical period of development, NR1 mRNA expression was lower in the contralateral AC, which received auditory signals from the plugged ear, than in the ipsilateral AC. However, no significant difference was observed between the rats deprived of hearing on P35 and those deprived of hearing on P42, the end of the critical period of auditory development. These results showed that monaural hearing deprivation during early postnatal development was associated with decreased NR1 mRNA expression in the contralateral AC and suggested the involvement of NR1 in auditory function during development. They also indicated that, during postnatal development, environmental factors changed the functional plasticity of neurons in the AC through NR1 receptor expression. Taken together, these findings provide a possible underlying mechanism for the development of postnatal auditory function.     

Distribution of glucocorticoid and mineralocorticoid receptor messenger RNA expression in human postmortem hippocampus

Corticosteroids bind to hippocampal glucocorticoid (GR) and mineralocorticoid (MR) receptors, thereby affecting behaviour and neurochemical transmission. Rat hippocampus has high levels of both receptors and their messenger RNAs (mRNA), but there is little information on receptors in human brain. We used in situ hybridization to determine the distribution of GR and MR mRNA expression in human hippocampus. Frozen sections of human postmortem hippocampus (5 patients, 58-88 years old, without cerebral pathology) were postfixed in paraformaldehyde and hybridized with 35S-UTP-labelled cRNA probes (transcribed in vitro from human cDNA subclones) under stringent conditions. Control included hybridization with sense probes and heterologous cRNA competition studies. GR mRNA was highly expressed in dentate gyrus, CA3 and CA4, but levels were significantly lower in CA1 and CA2. MR mRNA was also very highly expressed in hippocampus, with significantly higher levels in dentate gyrus and CA2, CA3 and CA4 than CA1. Controls confirmed the specificity of hybridization and there was little hybridization of sense probes. High GR and MR mRNA expression is found in both rat and human hippocampus but the subregional distributions clearly differ between the species.     

Differential synaptic localization of the glutamate transporter EAAC1 and glutamate receptor subunit GluR2 in the rat hippocampus

EAAC1, a neuron-specific glutamate transporter, is likely to play an important role in the regulation of glutamate levels in the synaptic cleft. Ultrastructural studies have demonstrated that the glutamate receptor subunit proteins (e.g., GluR2) are frequently preferentially located at the postsynaptic density of asymmetric synapses. While the glutamate/glutamate receptor interaction is likely to be influenced by the activity and location of the transporter molecules, the spatial localization of the transporter molecules relative to the receptor molecules is not well delineated. Thus, we analyzed the cellular, ultrastructural, and synaptic distribution of EAAC1 in the context of the distribution of the AMPA receptor subunit GluR2 in the hippocampus. While GluR2 and EAAC1 are both present in hippocampal projection neurons, their intracellular distribution patterns differ. Both GluR2 and EAAC1 are present in the dendritic membranes and cytoplasm; however EAAC1 has a distinctive punctate distribution in the dendrite compared to the more diffuse labeling reflected by GluR2. Pre-embedding ultrastructural studies also revealed cytoplasmic and membrane-associated pools of EAAC1 within dendritic shafts and spines, as well as in a subset of axonal profiles and terminals. Postembedding double label immunogold localization demonstrated a similar intraneuronal distribution, but in addition showed that membrane-associated EAAC1 is not intermingled with GluR2 within the synaptic complex, but in contrast is primarily located perisynaptically, often immediately outside the synaptic specialization. In addition, there is a significant presynaptic pool of EAAC1, whereas GluR2 is essentially absent from the pre-synaptic profile. Thus, membrane-associated EAAC1 within the synaptic region is ideally situated to restrict the site of action of glutamate with respect to ionotropic receptors to the synaptic cleft, as well as regulate glutamate levels in the perisynaptic and presynaptic domains, the ultrastructural sites that have been associated with metabotropic receptor localization.     

Maternal dexamethasone treatment in late gestation alters glucocorticoid and mineralocorticoid receptor mRNA in the fetal guinea pig brain

Development of the fetal hypothalamo-pituitary-adrenocortical (HPA) axis is critical for fetal maturation and responses to stress. Guinea pigs, unlike rats, give birth to mature young, and peak brain growth occurs around days 48-52 (75%) of gestation. There is extensive development of the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR) systems at the time of rapid brain growth in guinea pigs. Since approximately 10% of pregnant women are treated with synthetic glucocorticoids in late gestation, to promote fetal organ maturation, we tested the hypothesis that fetal exposure to glucocorticoids modifies developing GR and MR systems in the brain. Pregnant guinea pigs were subcutaneously injected with dexamethasone (dex; 1 mg/kg) or vehicle on days 50 and 51 of gestation (term=70 days). On day 52, guinea pigs were killed and the fetuses rapidly removed. Maternal dex treatment resulted in increased plasma cortisol concentrations in female fetuses, but decreased cortisol in male fetuses. Plasma thyroxine levels were increased in both female and male fetuses following maternal dex-treatment. Exposure to dex resulted in significant increases in MR and GR mRNA in the CA1-2 region of the hippocampus, and MR mRNA in the dentate gyrus in female fetuses. There was no effect of dex on GR or MR mRNA in the male fetuses. In conclusion, the effect of synthetic glucocorticoid on the developing brain GR and MR systems is sex-specific and is confined to very specific regions of the hippocampus. Since the hippocampus plays a central role in mediating glucocorticoid negative feedback of HPA function, alterations in the fetal development of corticosteroid receptors may form the basis of permanently modified HPA activity following fetal exposure to endogenous or synthetic glucocorticoid.     

Ionotropic glutamate receptor modulation preferentially affects NMDA receptor expression in rat hippocampus

Electrophysiological data suggest that alterations in the function of one glutamate receptor subtype may affect the function of other subtypes. Further, previous studies have demonstrated that NMDA receptor antagonists affect NMDA and kainate receptor expression in rat hippocampus. In order to address the mutual regulation of NMDA, AMPA, and kainate receptor expression in rat hippocampus, we conducted two experiments examining the effects of NMDA and non-NMDA glutamate receptor modulators on NMDA, AMPA, and kainate receptor expression using in situ hybridization and receptor autoradiography. NMDA receptor expression was preferentially affected by systemic treatments, as all drugs significantly altered [(3)H]MK-801 binding, and several drugs increased [(3)H]ifenprodil binding. GYKI52466 and aniracetam treatments resulted in changes in both [(3)H]ifenprodil binding and NR2B mRNA levels, consistent with the association of this subunit and binding site in vitro. There were more modest effects on AMPA and kainate receptor expression, even by direct antagonists. Together, these data suggest that ionotropic glutamate receptors interact at the level of expression. These data also suggest that drug regimens targeting one ionotropic glutamate receptor subtype may indirectly affect other subtypes, potentially producing unwanted side effects.     

Chronic neonatal nicotine upregulates heteromeric nicotinic acetylcholine receptor binding without change in subunit mRNA expression

Smoking during pregnancy chronically exposes the fetus to nicotine resulting in long-term behavioral and cognitive deficits. Nicotine binds to neuronal nicotinic acetylcholine receptors (nAChRs), pentameric ligand-gated ion channels widely expressed in the nervous system. Chronic nicotine upregulates high-affinity nAChRs in animals and smokers. Here we determined if chronic nicotine treatment during a developmental period corresponding to the human third trimester regulates nAChR expression. Rat pups were intubated orally three times per day with or without nicotine (6 mg/kg/day) from postnatal day 1 to 8. Subunit mRNA expression was assessed by in situ hybridization. Expression of heteromeric and homomeric nAChR receptor was evaluated by autoradiography using (125)I-epibatidine and (125)I-alphabungarotoxin, respectively. nAChR expression was analyzed in cortex, hippocampus, thalamus and medial habenula from autoradiograms using computer assisted image analysis. Nicotine induced significant upregulation of heteromeric but not homomeric nAChRs in hippocampus, cortex and thalamus without changes in subunit mRNA expression. No effect of chronic nicotine on receptor expression was detected in the medial habenula, suggesting that nicotine's effect was mainly on alpha4beta2-type heteromeric nAChRs. The nicotine-induced upregulation was reversed after nicotine withdrawal. Receptor blockade by DHbetaE, an antagonist for heteromeric alpha4/beta2 nAChRs, did not prevent upregulation but increased expression to a similar degree as nicotine. Combination of both drugs had a cumulative effect. Thus, although transient, intermittent nicotine exposure as seen in smoking mothers is sufficient to upregulate heteromeric nAChRs during a critical period of brain development and could contribute to the behavioral deficits found in children whose mother smoked.