Developmental changes in the expression of GABA_A receptor subunits (α₁, α₂, α₃) in the cat visual cortex and the effects of dark rearing

Abstract

The present study used Western blots and Northern slot blots to determine changes in the level of expression of GABA_A receptor subunits α₁, α₂, and α₃, in relation to the ‘critical period’ in cat visual cortex. Levels of the GABA_A α₁ subunit were lowest at 1 week, increased four-fold to a maximum at 10 weeks, and declined slightly (35%) into adulthood. Levels of the GABA_A α₂ and α₃ subunits were highest at 1 week of age, decreased two-fold by 10 weeks of age and were constant thereafter. Comparison between visual cortex from normal and dark-reared cats at 5 weeks and 20 weeks showed that α₁ and α₃ subunit expression was elevated in dark-reared animals by approximately 50% at both ages. α₂ expression was not affected. These results implicate the importance of a shift from putative immature to mature GABA_A receptor subunits during the critical period of visual cortex and in conjunction with parallel analysis of NMDA receptor subunit maturation, further support the notion that a changing excitatory/inhibitory balance is critical for neuronal plasticity.

Introduction

GABA_A receptors are ligand-gated chloride channels, which mediate the principal inhibitory neurotransmission in the adult brain. Structurally and functionally distinct GABA_A receptor subtypes have been identified and the heterogeneity is based on a family of at least 17 subunits that are further grouped into five major classes: α1–6, β1–4,γ1– 4, δ, ρ1–2 [4], [35], [42], [46], [48], [50]. The hetero-oligomeric protein complex of GABA_A receptor contains a pentameric structure as demonstrated by electron microscopic techniques [41]. As a result of different combinations of the 17 subunits, a large variety of GABA_A receptor subtypes are present in the central nervous system, although not all possible combinations are found [35], [42]. Differences in subunit composition are responsible for the functional diversity of GABA_A receptors in the brain.

Expression of GABA_A receptor subunits is regulated developmentally and shows differential regional and cellular distribution patterns [11], [12], [20], [23], [27], [29], [36]. These heterogeneities may explain why GABA mediates neurotrophic functions in the immature brain and synaptic inhibition in the adult brain [2], [32], [43]. The developmental regulation of GABA_A receptor subunits in brain is complex with different subunits showing different developmental time-courses of expression in different brain regions [43]. For the α subunit family, immunohistochemical and in situ hybridization studies indicate a shift from ‘immature’ (α₂ and α₃) to ‘mature’ (α₁) subunits during development. This switch has been documented in several species and brain regions and it may be temporally related to periods of synaptogenesis and enhanced functional plasticity [12], [18], [23], [27]. α₁ is the most ubiquitous α subunit and it is present in the major GABA_A receptor subtype found in adult brains [13], while α₂ and α₃ subunits are most common in developing brains [43].

The cat visual cortex is an ideal model to study development and plasticity of GABA_A receptor subunits because of its well-defined postnatal critical period for anatomical and physiological plasticity, as assessed by susceptibility to monocular deprivation. Plasticity in the visual cortex is absent until 3 weeks of age, peaks at about 5 weeks, declines to low levels at 20 weeks and disappears at about 1 year of age [8]. Rearing cats in total darkness alters the time-course of the critical period [7], [40]. The effect of darkness is to slow the entire course of the critical period [38] and thus it provides a means to isolate factors directly associated with critical period plasticity from those related to normal development. Several studies have indicated that GABA inhibition controls excitatory (NMDA) activity [28], [33] and that NMDA activation is involved in the regulation of GABA_A receptor subunit composition [15]. It has been proposed that the maturation of GABA inhibitory circuits acts as a ‘plasticity gate’ in postnatal critical period neuronal plasticity and is involved in the stabilization of mature cortical physiology [1], [9], [16], [19], [21], [24], [25]. The present study used Western and Northern blotting to determine: (1) developmental profiles of α₁, α₂ and α₃ subunit expression in normal cat visual cortex; and (2) whether visual input is necessary for developmental changes in GABA receptor subunit composition by comparing normal and dark reared 5 week (near the peak of the critical period) and 20 week (near the end of the critical period) cat visual cortex.