A study of the inhibitor of the crayfish neuromuscular junction by presynaptic voltage control. Vyshedskiy, Andrey and Jen-Wei Lin. Department of Biology, Boston University.
APStracts 3:0223N, 1996.
The inhibitor of the crayfish opener muscle was investigated by a presynaptic voltage control method. Two microelectrodes were inserted into the inhibitor and the amplitude and duration of presynaptic depolarization was controlled by a voltage clamp amplifier. Inhibitory postsynaptic potential (IPSP) was measured from a muscle fiber located near the presynaptic voltage electrode. Nonlinear summation of IPSP amplitudes was corrected after chloride equilibrium potential was measured. Using 5 ms presynaptic pulses, the depolarization-release coupling (D-R) curve constructed from IPSP peak amplitudes (IPSP cor ) had a threshold of about -35 mV and reached its maximal level at -5 to -10 mV. Depolarization beyond the maximum led to a suppression of neurotransmitter release. When transmitter release during a presynaptic pulse was completely suppressed, IPSPs activated by tail current could be identified with an average synaptic delay of 2.5 ms. Transmitter secretion triggered by a calcium current activated during the 5 ms pulses (IPSP on ) was also measured on the rising phase of an IPSP, at 2.5 ms after the end of the 5 ms pulses. D-R coupling plots measured from IPSP on exhibited a more pronounced suppression than that obtained from IPSP cor . The effect of presynaptic pulse duration on the level of transmitter release was analyzed. Transmitter release increased with increasing duration and was nearly saturated by 20 ms pulses depolarized to 0 mV. The following conditions were identified as necessary to obtain a consistent D-R curve with a clear suppression: (1) small animals, 3.8 cm head to tail, (2) 15C, (3) 40 mM TEA and 1 mM 4-AP, (4) an extracellular calcium concentration of 10 mM or less. In addition, a consistent correlation was found among the branching pattern of the inhibitor, the placement of the presynaptic electrode and the characteristics of the D-R curves. An ideal presynaptic electrode configuration involved placing the voltage electrode in a secondary branch, about 100 [mu] m from the main branch point, and placing the current electrode at the branch point. Postsynaptically, optimal recordings were obtained from muscle fibers innervated by a single branch of the inhibitor that originated from a point near the presynaptic voltage electrode. A cable-release model was constructed to evaluate the relationship between the shape of the D-R coupling curves and the space constants of the presynaptic terminals. A comparison between the model and the D-R coupling curves suggested that the space constant of an inhibitor branch on a muscle fiber is at least 8 times longer than its actual length. Therefore, the upper limit estimate of the space constant of a typical preparation is about 3 mm. Results reported here outline morphological and physiological conditions needed to achieve optimal control of the presynaptic branch of the crayfish inhibitor. The cable-release model quantitatively defines the extent of presynaptic voltage control.

Received 31 May 1996; accepted in final form 19 September 1996.
APS Manuscript Number J430-6.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 5 November 1996