An improved method for long-term measuring of hemolymph fluctuations of non-essential amino acids, GABA and histamine from freely moving crayfish.

The microdialysis method was adapted to obtain long-term hemolymph dialysates from the pericardial cavity of freely moving Procambarus clarkii crayfish, to measure fluctuations of non-essential amino acids, GABA and histamine by high-performance liquid chromatography using off-line fluorometric derivatization. Asp, Ala, Tau, GABA and histamine (HA) reached its maximal concentrations at the daybreak, whereas glutamate (Glu), Gln and Gly peaked at the end of the light period. The minimum and maximal detected amounts for each substance along the 24h cycle were (in microM): 20-300Asp, 100-200Glu, 400-700Gln, 400-600Gly, 100-200Tau, 150-300Ala, 2-10 GABA and 25-250HA.

Expression and functional characterization of GABA transporters in crayfish neurosecretory cells.

The effect of GABA on membrane potential and ionic currents of X-organ neurons isolated from the crayfish eyestalk was investigated. Under voltage-clamp conditions, GABA elicited an inward Na+ current followed by a sustained outward chloride current. Sodium current was partially blocked in a dose-dependent manner by antagonists of GABA plasma membrane transporters such as beta-alanine, nipecotic acid, 1-[2([(diphenylmethylene)imino]oxy)ethyl]-1,2,5,6-tetrahydro-3-pyridinecarboxylic acid hydrochloride (NO 711), and SKF89976-A at concentrations between 1 and 100 microm.