Gamma-aminobutyric acid (GABA) is the primary inhibitory neurotransmitter in the mammalian central nervous system, and most fast inhibitory neurotransmission in the brain is mediated by the GABAA receptors (GABARs). There are many families of GABAR subunits with multiple subtypes. The alpha subunit family is the largest, with six different subtypes (alpha1-alpha6). The expression of the subtypes is regulated regionally and developmentally and it is likely that each alpha subtype performs a unique physiological role. Subtype expression is also altered by pathological conditions, such as epilepsy. The subunit composition of the receptor affects many characteristics of the GABAR, but little is known about the effect of the identity of the alpha subtype on the channel's kinetic properties. The long-term goal of this work is to understand the functional significance of the structural heterogeneity of the GABAR and to determine which structural differences among the subunits underlie their different functional properties. The proposed work will examine this question by comparing the properties of recombinant receptors containing different alpha subtypes at the single channel, macropatch and whole-cell levels. The combination of these results will allow a complete description of the kinetic properties associated with each alpha subtype and may predict the behavior of synaptic receptors containing these subtypes. The structural differences responsible for distinct kinetic properties will be examined in detail for the alpha1and alpha 6 subtypes, which are the most functionally diverse in the alpha subunit family. To isolate the general structural domains responsible for the functional characteristics associated with each subtype, six chimeric alpha1 - alpha6 subunits will be examined. These chimeras were designed to isolate the functional contribution made from each of the four transmembrane domains. Once the general structural domains are isolated, the specific residues responsible for these characteristics will be identified through site-directed mutagenesis. The GABARs play an integral role in regulating neuronal activity. GABARs are important targets for drugs commonly used as sedatives, anxiolytics and anti-epileptics as well as for drugs of addiction, including barbiturates and alcohol. The activity of many of these modulators depends upon the alpha subtype composition of the receptor. The projects described will address the question of how structural differences among the alpha subtypes contribute to the functional properties of the channel. The results of this work will help us to understand how variations in the alpha subunit composition of native GABARs can affect the properties of neurons and their responses to GABA.
