Optical identification of calcium-dependent action potentials transiently expressed in the embryonic rat brainstem

Using multiple-site optical recording of transmembrane potential changes, we have found a new type of calcium-dependent action potential expressed transiently in the embryonic rat dorsal motor nucleus of the vagus nerve. Slice preparations with vagus nerve fibers attached were dissected from 12- to 16-day-old embryonic (E12-E16) rat brainstems, and they were stained with a voltage-sensitive merocyanine-rhodanine dye (NK2761). Electrical activities in response to vagal stimuli were optically recorded simultaneously from many sites using 1020- or 128-element photodiode array measuring systems. In brainstem preparations, two types of action potential-related optical signals were identified. One was detected from the dorsolateral region, and was related to sensory nerve activity (Type I). The other was detected from the dorsomedial region, and corresponded to the action potential in the dorsal motor nucleus of the vagus nerve (Type II). We found a difference in the ionic basis of the Type I vs Type II signals. The Type I signal was not altered in Ca2+-free bathing solution and was eliminated by tetrodotoxin, suggesting that the sensory nerve activity is mediated by Na+ currents. The Type II signal at early developmental stages (E12-E13, and some preparations in E14) was also independent of Ca2+. However, the Type II signal in later developmental stages (E15-E16, and some preparations in E14) did depend upon Ca2+: it was eliminated in Ca2+-free Ringer's solution, blocked by Cd2+, Ni2+ or Mn2+, and elicited in Sr2+-containing Ringer's solution, where CaCl2 was replaced with SrCl2. These results suggest that the cation which dominates the motoneuron action potential changes from Na+ to Ca2+ during development, and this change occurs around E14. With pharmacological analysis using Ca2+ channel blockers, we show that the Ca2+ channel mediating the motoneuron action potential is distinct from T-, L-, N-, P- or Q-type channels. Because the vagal action potential in adult mammals is mainly mediated by Na+, we suggest that a Ca2+ action potential mediated by a new type of Ca2+ channel is expressed transiently in the rat dorsal motor nucleus of the vagus nerve at particular stages of development.