Regulation of Serotonin_1A, Glucocorticoid, and Mineralocorticoid Receptor in Rat and Human Hippocampus: Implications for the Neurobiology of Depression

Abstract

Background: Disturbances of the limbic–hypothalamic–pituitary–adrenal axis and the serotonin system are commonly found in depressive illness. Studying the effect of stress on these two neurobiological systems may give us important clues into the pathophysiology of affective illness and help us understand how stress and mood disorders are related.

Methods: We studied the effect of chronic unpredictable stress and antidepressant treatment on serotonin 1A (5-HT_1A), glucocorticoid (GR), and mineralocorticoid (MR) receptor levels in rat hippocampus, using in situ hybridization and receptor autoradiography. We also used in situ hybridization to quantify hippocampal 5-HT_1A, GR, and MR messenger (mRNA) levels in a small group of suicide victims with a history of depression, compared to matched controls (n = 6).

Results: We found that rats subjected to chronic unpredictable stress showed a significant elevation of basal plasma corticosterone compared to nonstressed rats. Chronic stress also caused a decrease in 5-HT_1A mRNA and binding in the hippocampus. In addition, chronic stress produced alterations on the MR/GR mRNA ratio in this same region. The decreases in 5-HT_1A mRNA and binding, as well as the MR/GR alterations, were prevented in animals that received imipramine or desipramine antidepressant treatment. Zimelidine was unable to reverse stress-induced increases in corticosterone, and was only partially successful in preventing the stress-induced receptor changes in the hippocampus. Suicide victims with a history of depression showed changes that were very similar to the changes found in chronic stress.

Conclusions: Alterations in hippocampal 5-HT_1A levels and in the MR/GR balance may be one of the mechanisms by which stress may trigger and/or maintain depressive episodes.

Introduction

Research studies have implicated disturbances in the serotonin (5-HT) system Meltzer 1988, Meltzer 1989and the limbic–hypothalamic–pituitary–adrenal (LHPA) axis Gold et al 1988; Kathol et al 1989as the neurobiological “alterations” most consistently associated with affective illness. Although abnormalities in these two systems are usually studied individually, their interaction in the brain, as it relates to the pathophysiology of depression, has not been as extensively studied.

In reviewing the clinical, psychological, and biological literature on depressive illness, one factor that emerges as being closely associated with depression is stress. Stress and depression have been linked in a variety of ways; for example, both physical and psychological stressors have been shown to be temporally (and perhaps causally) related to the onset of depressive episodes Post 1992. Some studies have suggested that, at least for recurrent depression, stressful life events are more common in “nonendogenous depression” Frank et al 1994. Other studies have found that stressful life events are significantly correlated even with the first episode of psychotic/endogenous depression Brown et al 1994. Another important link between depression and stress is the fact that both the LHPA and 5-HT systems, in addition to being involved in the pathophysiology of depression, are also critical contributors to the neurobiology of stress McEwen 1987. Therefore, studying the neurobiology of stress by focusing on these two systems may give us important clues into the pathophysiology of affective illness, shed light on the actions of antidepressants, and begin to reveal how stress and mood disorders are related.

The LHPA axis is a neuroendocrine system that is closely linked to stress in mammals. This system is geared to allow a swift response to stressful stimuli and ultimately a return to homeostasis, through complex feedback mechanisms. The parvocellular neurons of the paraventricular nucleus (PVN) in the hypothalamus represent the final common path for the integration of the stress response in the brain. They express corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP), both of which are secretagogues of the LHPA. In response to physiological and psychological stressors, CRH and AVP are secreted into the hypophyseal portal circulation to reach the anterior pituitary, where they synergistically stimulate the proopiomelanocortin (POMC)-producing cells of the anterior lobe to release adrenocorticotropic hormone (ACTH), an end product of POMC precursor molecule processing. ACTH transported in the circulation interacts with the adrenal, causing elevation of plasma glucocorticoids. Repeated exposure to elevated glucocorticoid levels has been implicated in neuronal death, among other deleterious effects. Therefore, inhibition of steroid secretion is also an important component of this system. This inhibition is partly achieved by glucocorticoid binding to specific corticoid receptors in the pituitary and in limbic structures Lopez et al 1991.

Two types of corticoid receptors have been described in the brain based on biochemical and functional characteristics McEwen 1991. Type I or mineralocorticoid receptor (MR) resembles the kidney mineralocorticoid receptor and has stringent specificity, binding selectively corticosterone, the main glucocorticoid of the rat. In the brain, MR is most densely localized in hippocampal and septal neurons and is described as a “high-affinity, low-capacity corticoid receptor system.” Conversely, type II or glucocorticoid receptor (GR) is known as a “low-affinity, high-capacity receptor system.” GR receptors are widely distributed in brain, including hippocampus, hypothalamus, and pituitary cells; however, they bind corticosterone with a lower affinity compared to MR. Their highest affinity is for potent synthetic glucocorticoids, such as dexamethasone McEwen 1991. These receptor characteristics complement each other and put the MR and GR in a position to modulate LHPA responses. The MR receptors appear to be operative at low corticosterone concentrations and may offer tonic inhibition to the axis during the nadir of the circadian rhythm De Kloet et al 1988; McEwen 1991. When high concentrations are present, MR receptors saturate, and the GR receptors appear to ensure the return of homeostasis. It is thus apparent that dysfunction on any of the complex pathways within the system could severely affect the organism’s adaptive capacity and expose the brain to high glucocorticoid concentrations.

Hyperglucocorticoid states have been of interest to psychiatric research both as the consequence and the cause of depressive symptoms. Studies that investigate glucocorticoid hypersecretion as a consequence of depression have focused on the fact that a significant proportion of depressed patients show dysregulation of the LHPA axis. This dysregulation is manifested by cortisol hypersecretion, failure to suppress cortisol secretion after dexamethasone administration, and blunted ACTH response to CRH administration Carroll et al 1976; Gold et al 1986; Kathol et al 1989; Sachar et al 1973. Although these findings are concerned with peripheral measures of LHPA activity, there is also good evidence of increased central drive, based on increased activity at the nadir of the circadian rhythm Young et al 1995, impaired negative feedback Young et al 1991, as well as more direct findings of elevated CRH in the cerebrospinal fluid (CSF) of depressed patients Nemeroff et al 1984. These LHPA alterations are thought to reflect either a central neurotransmitter abnormality, the stress of the illness, or a combination of both.

Studies that investigate hyperglucocorticoid states as a cause for depressive illness are less numerous, and have been stimulated by the fact that the final products of the LHPA axis (cortisol in humans, corticosterone in rats) have been shown to have profound effects on mood and behavior McEwen 1987. For example, a high incidence of depression is linked to pathologies involving elevated corticosteroid levels, such as Cushing’s syndrome. This corticosteroid-induced depression usually disappears when corticosteroid levels return to normal Kathol 1985; Murphy 1991.

It is clear, from both animal and clinical studies Kathol 1985; McEwen 1987; Murphy 1991, that circulating steroid levels provide important hormonal control of affect, which may be mediated by steroid-induced modulation of central limbic circuitry. The precise mechanism by which corticosteroids exert this influence on affect is not well understood; however, this mechanism is likely to involve interactions with brain neurotransmitters, since we know that central control of affect is intimately associated with the actions of the monoaminergic molecules 5-HT, norepinephrine, and dopamine. Effective clinical treatment of affective disorders such as depressive illness has been accomplished with drugs that act to potentiate the action of these monoaminergic neurotransmitter systems Baker and Greenshaw 1988. For 5-HT in particular, a new generation of specific agents with strong antidepressant properties has been developed Lucki 1991; consequently, alterations in serotonergic circuitry has been proposed as one of the central components of affective disease. Thus, it is likely that functional interactions between corticosteroids and the central serotonergic system may be significant, providing a biochemical basis for hormonally induced alterations in central nervous system (CNS) circuits related to emotional control.

Animal studies have in fact shown that corticosteroids can alter several elements of serotonergic neurotransmission. Removal of circulating corticosteroids by adrenalectomy (ADX) results in anatomically specific decreases in indices of serotonin metabolism, while stressful procedures, which raise corticosteroid levels, produce corresponding increases in serotonin turnover Curzon et al 1972; Van Loon et al 1981; however, corticosteroids may also act to directly modulate serotonergic neurotransmission by regulating 5-HT receptors. Autoradiographic studies Biegon et al 1985first identified increased 5-HT₁ receptor binding in the rat hippocampal formation 1 week after bilateral adrenalectomy. Subsequent investigations confirmed the sensitivity of 5-HT₁ receptors to circulating corticosteroid levels De Kloet et al 1986and indicate that specific hippocampal subfields are exquisitely sensitive to adrenal steroids. More recent electrophysiological studies have shown a suppression of 5-HT-induced hyperpolarization within Ammon’s horn 1 (CA1) pyramidal cells after brief application of steroids Joels et al 1991, establishing a functional coupling for steroid–serotonin receptor interactions within the hippocampus. It, therefore, seems likely that the high concentration of corticosteroid receptors within the hippocampus underlies the sensitivity of 5-HT receptors to corticosteroid regulation in this region. It is also well established that the hippocampus is a central component of limbic circuitry and is fundamental in controlling aspects of cognitive and behavioral functions. The ascending serotonergic innervation of hippocampal neurons arising in midbrain raphe nuclei provides one means by which the serotonergic system may act to regulate limbic function. Thus, the hippocampus represents a key anatomical structure involved in the central control of LHPA axis function and limbic circuitry. As such, this area provides a unique anatomical environment in which to study the molecular interplay between serotonergic systems and corticosteroids.

The actions of 5-HT are mediated by multiple receptors. Serotonin receptors were originally divided into two subtypes, 5-HT₁ and 5-HT₂, on the basis of pharmacologic profile Peroutka and Snyder 1979; however, more recent pharmacologic and biochemical data indicate the presence of at least six major 5-HT receptor families or classes: 5-HT₁ through 5-HT₇, with some families containing multiple subtypes Hoyer and Martin 1996. Thus, the 5-HT₁ receptor family, defined as exhibiting high affinity for 5-HT, has been subdivided into five receptor subtypes, 5-HT_1A–1F. (The 5-HT_1c receptor was reclassified as 5-HT_2c, due to its resemblance to members of the 5-HT₂ family.)

The 5-HT_1A receptor in particular has been identified both as an inhibitory somatodendritic receptor in raphe serotonergic cells and a postsynaptic receptor in selective serotonergic terminal fields Hall et al 1985. Its abundance in the limbic system, as well as evidence from animal and clinical studies, strongly suggest that the 5-HT_1A receptor plays an important role in the pathophysiology of mood disorders. Some investigators have hypothesized that a “balance” between the 5-HT_1A and the 5-HT_2A receptors is essential to antidepressant action. This is based on the observation that, while the net effect of antidepressant administration seems to be functional down-regulation of 5-HT_2A receptors in the neocortex, the opposite is often observed for 5-HT_1A receptors in the hippocampus. For example, antidepressant drugs have been found to up-regulate hippocampal 5-HT_1A receptor electrophysiological responsiveness Blier et al 1988and possibly increase receptor number Welner et al 1989, although this last observation is not universal Watanabe et al 1993. This suggests that this receptor subtype may be important in relation to the therapeutic actions of these compounds. In humans, selective 5-HT_1A agonists, such as gepirone and buspirone, can effectively treat generalized anxiety disorder Rickels 1990as well as depression Fabre 1990, including the melancholic subtype Robinson et al 1990. Buspirone has also been reported to augment the antidepressant response in patients who are antidepressant nonresponders Jacobsen 1991.

5-HT_1A is the most abundant serotonin receptor in the hippocampus, where it is colocalized with glucocorticoid and mineralocorticoid receptors, making this structure an ideal anatomical substrate for studying steroid–5-HT_1A receptor interactions. Several investigators have reported increases in 5-HT_1A receptor binding and gene expression in response to ADX Chalmers et al 1994; Mendelson and McEwen 1990, demonstrating that this receptor subtype is under tonic inhibition by corticosterone. Thus, it is possible that the mechanisms underlying corticosteroid regulation of 5-HT_1A receptors may contribute to steroid-mediated modulation of affective state and, as such, may represent an important linkage both to the pathophysiology of affective disorders and to psychotherapeutic drug action.

The present paper reports previously unpublished series of preclinical experiments in the rodent, designed to further our understanding of the mechanisms involved in corticosteroid-mediated modulation of the 5-HT_1A receptor at the molecular and neuroanatomical level under conditions related to chronic stress and pathophysiological levels of steroids. Specifically, we have used a chronic unpredictable stress model to ask: 1) How do hippocampal 5-HT_1A receptors react to endogenously elevated corticosteroids? 2) Can antidepressant treatment alter the 5-HT_1A receptor under conditions of chronic stress? 3) What is the effect of a milder stress (swim), which produces only modest elevations of corticosterone? 4) Can we modify the effects of chronic stress on hippocampal 5-HT_1A receptors by preventing the increase in corticosterone, using adrenalectomy? 5) Are there differences between a specific serotonin reuptake inhibitor (SSRI) and a noradrenergic reuptake inhibitor in their ability to modify the effects of chronic stress? 6) What are the roles of MR and GR in mediating hippocampal 5-HT_1A receptor regulation? We also report results from a postmortem study, investigating the regulation of 5-HT_1A, MR, and GR in the hippocampus of suicide victims with a history of major depressive disorder. Here we ask: Will we find evidence of 5-HT_1A, GR, and/or MR dysregulation that parallels the abnormalities found in models of chronic stress? In this way we have identified a molecular linkage between the LHPA axis and the 5-HT system and discussed the potential relevance of these to the mechanism of action of antidepressant drugs and the pathophysiology of affective disorders.