Localization of glutamate and glutamate transporters in the sensory neurons of Aplysia

The sensorimotor synapse of Aplysia has been used extensively to study the cellular and molecular basis for learning and memory. Recent physiologic st udies suggest that glutamate may be the excitatory neurotransmitter used by the sensory neurons (Dale and Kandel [1993] Proc Natl Acad Sci USA. 90:716 3-7167; Armitage and Siegelbaum [1998] J Neurosci. 18: 8770-8779). We furth er investigated the hypothesis that glutamate is the excitatory neurotransm itter at this synapse. The somata of sensory neurons in the pleural ganglia showed strong glutamate immunoreactivity. Very intense glutamate immunorea ctivity was present in fibers within the neuropil and pleural-pedal connect ive. Localization of amino acids metabolically related to glutamate was als o investigated. Moderate aspartate and glutamine immunoreactivity was prese nt in somata of sensory neurons, but only weak labeling for aspartate and g lutamine was present in the neuropil or pleural-pedal connective. In cultur ed sensory neurons, glutamate immunoreactivity was strong in the somata and processes and was very intense in varicosities; consistent with localizati on of glutamate in sensory neurons in the intact pleural-pedal ganglion. Cu ltured sensory neurons showed only weak labeling for aspartate and glutamin e. Little or no gamma-aminobutyric acid or glycine immunoreactivity was obs erved in the pleural-pedal ganglia or in cultured sensory neurons. To furth er test the hypothesis that the sensory neurons use glutamate as a transmit ter, in situ hybridization was performed by using a partial cDNA clone of a putative Aplysia high-affinity glutamate transporter. The sensory neurons, as well as a subset of glia, expressed this mRNA. Known glutamatergic moto r neurons B3 and B6 of the buccal ganglion also appeared to express this mR NA. These results, in addition to previous physiological studies (Dale and Kandel [1993] Proc Natl Acad Sci USA. 90:7163-7167; Trudeau and Castellucci [1993] J Neurophysiol. 70:1221-1230; Armitage and Siegelbaum [1998] J Neur osci. 18:8770-8779)) establish glutamate as an excitatory neurotransmitter of the sensorimotor synapse. (C) 2000 Wiley-Liss, Tnc.