Suggestion of a role for calmodulin and phosphorylation in regulation of rabbit ileal electrolyte transport: Effects of promethazine

Suggestion of a role for protein phosphorylation in the regulation of intestinal active NaCl transport was found by studying the effects of low concentrations of promethazine on Ca²⁺-calmodulin (CaM)-dependent protein phosphorylation of ileal microvillus membranes and on active ileal electrolyte transport. Ca²⁺-CaM increased the phosphorylation of six microvillus peptides (M(r) 137,000, 116,000, 77,000, 58,000, 53,000, and 50,000) in a concentration-dependent manner. Promethazine inhibited the Ca²⁺-CaM-induced increases in each of these phosphorylations. The effect of promethazine was concentration dependent, with concentrations of 5-12 μM (mean 8μM) causing 50% inhibition. Promethazine also caused a concentration-dependent increase in net Cl absorption and decrease in the ileal short-circuit current, with 9 μM promethazine causing a change in short-circuit current 50% of maximum. The promethazine effect on microvillus membrane phosphorylation was specific, since neither cAMP- and cGMP-induced phosphorylation in the microvillus membrane nor the stimulation by Ca²⁺-CaM of myosin light chain kinase phosphorylation of myosin light chain were affected by promethazine. The similar, and unusual sensitivity to low concentrations of promethazine on ileal microvillus membrane phosphorylation increased by Ca²⁺-CaM and on ileal electrolyte transport is consistent with Ca²⁺-CaM-dependent microvillus membrane phosphorylation being involved in the regulation of active electrolyte transport in ileal absorptive cells.