Elsevier

Brain Research

Volumes 1073–1074, 16 February 2006, Pages 120-130
Brain Research

Research Report
Age-related alterations in GABAA receptor subunits in the nonhuman primate hippocampus

https://doi.org/10.1016/j.brainres.2005.12.036Get rights and content

Abstract

Pharmacological studies have documented that altered drug responses, particularly to benzodiazepines, are common in elderly populations. While numerous factors may contribute to changes in drug response, age-related alterations in the molecular composition of GABAA receptors may be a key factor in regulating these responses. We employed quantitative densitometry to examine the cytological features and density of highly prevalent hippocampal GABAA receptor subunits (α1 and β2/3) in young and aged rhesus monkeys. α1 and β2/3 subunit immunostaining was differentially distributed throughout the hippocampus. In addition, β2/3 immunolabeling in aged monkeys was characterized by marked intersubject variability in labeling intensity, with dramatic reductions present in 3 of 5 samples. α1 immunolabeling in aged monkeys was significantly reduced in the CA2 and CA3 subregions, and in hilus/polymorphic layer of the dentate gyrus. Collectively, our findings demonstrate that not only are GABAA receptor subunits differentially distributed throughout the hippocampus, but they are also differentially altered with increased age—changes that may have an important impact on the binding properties of GABAA receptor pharmacological agents.

Introduction

Gamma-aminobutyric acid (GABA), the major inhibitory neurotransmitter in the mammalian CNS, hyperpolarizes neuronal membranes by opening a Cl channel intrinsic to the GABAA receptor. Molecular cloning studies have established that genes encoding subunits of the GABAA receptor constitute a large multigene family (Olsen and Tobin, 1990, Whiting et al., 1999 for review). These subunits are grouped according to sequence identity: α1–6, β1–4, γ1–4, ρ1–3, δ, ε, π, and θ (Olsen and Tobin, 1990, MacDonald and Olsen, 1994, Rabow et al., 1995, Sieghart, 1995, Mohler et al., 1996, Barnard et al., 1998, Bonnert et al., 1999, Whiting et al., 1999). The α and β subunit families are among the most studied of the GABAA receptor subunits. While the α family of subunits is involved in conferring sensitivity to benzodiazepines, the β family of subunits is sensitive to the actions of muscimol and other allosteric agonists (see Paul, 1995 for review). As will be further explored in the discussion, the presence of the GABA α1 subunit yields a receptor with a much higher affinity for GABA binding, which can be dramatically altered by benzodiazepine binding (Luddens and Wisden, 1991).

The α1 and β2/3 subunits are among the most widely expressed GABAA receptor subunits in the adult human brain. While the β subunits are not directly involved in benzodiazepine binding (see Paul, 1995 for review), their presence may play a role in regulating benzodiazepine affinity (Pritchett et al., 1988). β subunits also contain consensus sequences for phosphorylation (by protein kinase A), a factor which may result in receptor desensitization (Paul, 1995). Similar to the α1 subunit, given their prevalence, the β2/3 subunits likely play an important role in altered GABA receptor function in elderly populations.

While it is well established that embryonic and early postnatal GABAA receptors in the rat brain greatly differ from those in the adult, reports on altered GABAergic function during aging are less well established. However, an investigation of age-related alterations in GABAA receptors is central to understanding altered drug sensitivities in the elderly (Meyer, 1992, Monane, 1992).

In the CNS, GABAA receptor subunits are differentially expressed within neurons and glia across the neuroaxis. In situ hybridization studies have demonstrated that although considerable overlap exists, each subunit displays a unique pattern of expression in the adult rodent brain (Laurie et al., 1992a, Laurie et al., 1992b, Persohn et al., 1992, Wisden et al., 1992). For example, the most abundant mRNAs in rat CA3 pyramidal neurons correspond to the α2, α5, β1, β3, and γ2 receptor subunits (Wisden et al., 1992). Lower amounts of the α1 and α4 mRNAs are also present. Moreover, the expression patterns of GABAA receptor subunits change temporally during embryonic and postnatal development, as well as in the aging brain (Laurie et al., 1992b). The patterns of mRNA expression have been confirmed at the protein level using subunit-specific antibodies. Studies of cultured hippocampal neurons show temporal changes with maturity with respect to the receptor subunits expressed (Killisch et al., 1991). Immunohistochemical studies of the human hippocampus have also confirmed overlapping, yet unique patterns of immunolabeling (Houser et al., 1988), particularly for β2/3 and α1 (Mizukami et al., 1997, Mizukami et al., 1998a).

In the present study, we sought to determine whether specific subunits of the hippocampal GABAA receptor undergo age-related alterations in the rhesus monkey. The hippocampus was chosen for study, in part, because previous investigations have demonstrated alterations in α1 and β2/3 protein and mRNA within the hippocampus of patients with Alzheimer's disease (AD) (Mizukami et al., 1997, Mizukami et al., 1998a, Mizukami et al., 1998b).

To date, few studies have examined the patterns of expression of GABAA receptor subunits in the aging primate brain. The use of nonhuman primates is advantageous in that the potentially confounding issues related to postmortem interval and agonal state can be avoided. In addition, a comparison of the data obtained through studies of the aged nonhuman primate brain and AD patients is central to determining which alterations in GABAA receptor subunits are associated with normal aging, and which represent a neuropathologic process.

Section snippets

GABAA β2/3 in young monkeys

Following low magnification examination of young subjects labeled for β2/3, we observed intense peroxidase reaction product within the molecular layer of the dentate gyrus, as well as in the CA1 subregion (Figs. 1A–D). Less intense, but still conspicuous immunolabeling was observed in the CA2, CA3, and hilar subfields. Upon examination with high magnification optics, we observed very intense immunolabeling within both the inner and outer segments of the DG molecular layer (Fig. 1B). Within the

Discussion

The present study represents the first detailed immunohistochemical study of GABAA receptor subunits β2/3 and α1 in the hippocampus of the young and aged rhesus monkey. The results obtained from young monkeys provide important baseline information on the chemoarchitecture of the GABAA receptor in the monkey hippocampus. Moreover, our findings suggest that GABAA receptor subunits undergo regionally selective alterations in density and morphologic characteristics with increasing age.

Consistent

Animals

Chinese rhesus monkeys (Macaca mulatta) were wild-caught at birth and brought into the United States as part of a group-housed breeding colony until the age of 18–20. Animals were then individually housed and aged without experimentation at the Army Primate Facility in Texas. Aged monkeys were then purchased by Rush University Medical Center where they were individually housed at the Biological Resources laboratory at the University of Illinois, Chicago. Housing and handling of animals were

Acknowledgment

Grant Sponsor: NIH AG-15472.

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