No abstract

No abstract

No abstract

In 1957, Bazemore, Elliott, and Florey' showed that y-aminobutyric acid (GABA) blocked the discharges of crayfish stretch-receptor neurons. This finding focused attention on the possible role of GABA as an inhibitory transmitter compound. At various crustacean synapses GABA was found to mimic the effects of inhibitory (I) nerve stimulation.2-9 Furthermore, in the lobster nervous system GABA is highly concentrated in I neurons, the ratio of GABA concentrations in I and excitatory (E) axons being more than 100:1.10-12 Although this evidence is consistent with GABA being an inhibitory transmitter compound, a crucial experiment has hitherto been lacking: the demonstration that GABA is released from I-nerve terminals in response to nerve stimulation.The results of the present study show that stimulation of the I-nerve innervating various lobster muscles causes a release of GABA into the bathing fluid; the amount released is proportional to the number of stimuli applied.Materials and Methods.-Saline medium used to wash nerve-muscle preparations had the following composition: 460 mM NaCl, 15.6 mM KC1, 26 mM CaCl2, 8.3 mM MgSO4.Dowex-50W-X-2, H+ resin (100-200 mesh) was obtained from Baker Chemical Co. After fine particles were removed by repeated decantation, the resin was cycled through ammonium and hydrogen forms, and was freshly washed with 5-bed vol of 2 N hydrochloric acid and 2-bed vol of water immediately before use.Dowex-l-X2, Cl-resin (100-200 mesh) was obtained from Bio Rad Laboratories. The resin was thoroughly washed with 2 N hydrochloric acid, and then with 2 N acetic acid to convert it to the acetate form; resin columns were freshly washed with 5-bed vol of 2 N acetic acid and 5-bed vol of water immediately before use.3-H3-GABA (specific activity 5 c/mmole) was synthesized from DL-3-H3-glutamate (New England Nuclear Corp., Boston, Mass.) by decarboxylation with a bacterial glutamate decarboxylase.Nerve-muscle preparations: Opener muscles from the crusher and cutter claws of 0.5-kg lobsters (Homarus americanus) were set up as illustrated in Figure 1. The single E and I axons innervating the muscle lie in separate bundles in the carpopodite segment: these were dissected and stimulated separately with suction electrodes. The closer muscle and overlying connective tissue were carefully removed to expose the opener muscle. The opener muscles remained in situ in the exoskeleton which formed a convenient chamber for washing with saline medium. Junctional (synaptic) potentials were recorded intracellularly from muscle fibers with a glass microelectrode filled with 2 M potassium citrate. Chilled, oxygenated saline medium was pumped onto the proximal end of the muscle at a rate of approximately 2 ml/min. The perfusion medium percolated through the muscle and dripped out of the cut end of the dactyl (cleaned of internal tissue). The temperature of the effluent medium was 12-14°. I or E nerves were stimulated at frequencies of 5-20 impulses/sec for the first 15 min of the collection period.Superficial flexor muscles ...

No abstract