Voltage-dependent sodium and calcium currents of rat myenteric neurons in cell culture

Abstract

1. Inward currents of myenteric neurons that had been grown in cell cultures prepared from the small intestines of neonatal or young adult rats were studied with tight seal whole-cell recordings. The kinetic and pharmacological properties of these neurons were analyzed. 2. All neurons had rapidly inactivating, tetrodotoxin (TTX)-sensitive Na+ currents that could be evoked by steps to potentials more positive than -50 mV. Holding potentials more negative than -65 mV were necessary to remove steady-state inactivation. No TTX-insensitive Na+ currents were observed, thus the ability of subsets of myenteric neurons to fire action potentials in TTX must depend upon their density of Ca2+ channels. 3. Ca2+ and Ba2+ currents were studied in neurons perfused internally with CsCl to block K+ currents and bathed with solutions containing TTX and antagonists of K+ channels. Currents were significantly larger when Ba2+ replaced Ca2+ as the charge carrier. Cd2+ and Gd3+ blocked Ca2+ and Ba2+ currents rapidly and reversibly. High-voltage-activated (HVA) Ca2+ and Ba2+ currents were observed in all neurons. Too few neurons possessed detectable low-voltage-activated Ca2+ currents to permit detailed study. 4. HVA Ca2+ and Ba2+ currents evoked from holding potentials more negative than -50 mV could be divided into two kinetically distinguishable components with very different rates of inactivation. A "decaying" component inactivated relatively rapidly with a t1/2 of 25-75 ms. A "sustained" component inactivated quite slowly with a t1/2 of 1-5 s. At more positive holding potentials, only the sustained component was observed. Although the two kinetically distinguishable components had different current-voltage relationships, they had indistinguishable rates of deactivation: a single time constant was sufficient to fit the decay of tail currents. The relative amplitudes of the two components varied considerably among different neurons. 5. Ca2+ and Ba2+ currents could be divided into two pharmacologically distinct components on the basis of sensitivity to omega-conotoxin GVIA (I omega CgTX) and to dihydropyridine antagonists (IDHP). At holding potentials more positive than -70 mV, a combination of omega CgTX and DHPs completely blocked Ca2+ and Ba2+ currents in most neurons. At holding potentials more negative than -50 mV, I omega CgTX and IDHP each contained decaying and sustained components. I omega CgTX activated more slowly than did IDHP. The DHP agonist Bay K8644 increased the amplitude of IDHP and slowed its rate of deactivation. 6. The results suggest that myenteric neurons may have as few as two subtypes of HVA Ca2+ channels; omega CgTX-sensitive ones and DHP-sensitive ones.(ABSTRACT TRUNCATED AT 400 WORDS)