Effects of temperature on the experimental reliability of the isolated rat phrenic nerve/diaphragm preparation.

The isolated rat phrenic nerve/diaphragm preparation has proven to be a useful tool for examining neuromuscular transmission and function in vitro. This preparation allows for the simultaneous examination of direct phrenic nerve stimulation as well as direct muscle stimulation. While the isolated phrenic nerve/diaphragm has been extensively utilized (Bulbring, 1946; Tsai, 1985; Wali et al., 1988), the effects of isolation temperature on the performance of this in vitro model have not been reported. Cooling of the phrenic nerve (to OOC) in anesthetized dogs in vivo produces “a quick increase in phrenic conduction time and a decrease in diaphragmatic muscle action potential leading to a complete nerve conduction block” (Dureuil et al., 1987). We report that cooling of the rat phrenic nerve/diaphragm preparation to ambient (room) temperature after isolation in vitro produces a poor success rate of the model. By immediate placement of the isolated tissues into warm (37°C) buffer instead of ambient temperature buffer, the reliability and success rate of the preparation increases considerably. The poor success rate at ambient temperature could be due to irreversible nerve conduction block, as was described by Dureuil et al. (19871, to subtle changes in the responsiveness of the muscle itself, or to changes in the quanta1 release of transmitter (acetylcholine), which has been previously demonstrated by Datyner and Gage (1980).