No abstract

The mutations responsible for several human neurodegenerative disorders are expansions of translated CAG repeats beyond a normal size range. To address the role of repeat context, we have introduced a 146-unit CAG repeat into the mouse hypoxanthine phosphoribosyltransferase gene (Hprt). Mutant mice express a form of the HPRT protein that contains a long polyglutamine repeat. These mice develop a phenotype similar to the human translated CAG repeat disorders. Repeat containing mice show a late onset neurological phenotype that progresses to premature death. Neuronal intranuclear inclusions are present in affected mice. Our results show that CAG repeats do not need to be located within one of the classic repeat disorder genes to have a neurotoxic effect.

Syntaxin 1A inhibits GABA uptake of an endogenous GABA transporter in neuronal cultures from rat hippocampus and in reconstitution systems expressing the cloned rat brain GABA transporter GAT1. Evidence of interactions between syntaxin 1A and GAT1 comes from three experimental approaches: botulinum toxin cleavage of syntaxin 1A, syntaxin 1A antisense treatments, and coimmunoprecipitation of a complex containing GAT1 and syntaxin 1A. Protein kinase C (PKC), shown previously to modulate GABA transporter function, exerts its modulatory effects by regulating the availability of syntaxin 1A to interact with the transporter, and a transporter mutant that fails to interact with syntaxin 1A is not regulated by PKC. These results suggest a new target for regulation by syntaxin 1A and a novel mechanism for controlling the machinery involved in both neurotransmitter release and reuptake.

␥-Aminobutyric acid (GABA) transporters on neurons and glia at or near the synapse function to remove GABA from the synaptic cleft. Recent evidence suggests that GABA transporter function can be regulated, although the initial triggers for such regulation are not known. One hypothesis is that transporter function is modulated by extracellular GABA concentration, thus providing a feedback mechanism for the control of neurotransmitter levels at the synapse. To test this hypothesis, GABA uptake assays were performed on primary dissociated rat hippocampal cultures that endogenously express GABA transporters and on mammalian cells stably expressing the cloned rat brain GABA transporter GAT1. In both experimental systems, extracellular GABA induces chronic changes in GABA transport that occur in a dose-dependent and time-dependent manner. In addition to GABA, ACHC and nipecotic acid, both substrates of GAT1, up-regulate transport; GAT1 transport inhibitors that are not transporter substrates down-regulate transport. These changes occur in the presence of blockers of both GABA A and GABA B receptors, occur in the presence of protein synthesis inhibitors, and are not influenced by intracellular GABA. Surface biotinylation experiments reveal that the increase in transport is correlated with an increase in surface transporter expression. This increase in surface expression is due, at least in part, to a slowing of GAT1 internalization in the presence of extracellular GABA. These data suggest that the GABA transporter fine-tunes its function in response to extracellular GABA and would act to maintain a constant level of neurotransmitter at the synaptic cleft. GABA1 transporters are members of a large family of Na ϩ -dependent neurotransmitter reuptake proteins, located on the plasma membrane of neurons and glia, that function in part to determine neurotransmitter levels in the synaptic cleft (1). Demonstration of a physiological role for GABA transporters comes from experiments involving specific GABA uptake inhibitors; these inhibitors prolong the decay phase of GABA A receptor-mediated post-synaptic potentials (2) and both prolong the decay phase and increase the magnitude of responses mediated by the G protein-coupled GABA B receptor (2-4). GABA transporters also play a physiological role in toad and catfish horizontal cells where calcium-independent GABA efflux through the transporter is a principal mode of neurotransmitter release (5). GABA transporters also have a pathophysiological role. There is decreased calcium-independent GABA release in the affected hippocampus of temporal lobe epileptics. This decrease in transporter-mediated efflux is correlated with fewer GABA transporters and is hypothesized to result in decreased inhibitory tone (6).Not only can GABA transporters regulate neuronal signaling, transport itself can be regulated. This is true for GABA transporters and other members of this family as well (for review see Refs. 7 and 8). Functional modulation occurs through a variety of second messengers such as...