Pharmacomechanical coupling in rat vas deferens: Effects of agents that modulate intracellular release of calcium and protein kinase C activation.

The effects of agents that modulate intracellular release of calcium and pr otein kinase C (PKC) activation on noradrenaline (NA)-induced contractions of epididymal vas deferens in calcium-free/EGTA (1 mM) medium were investig ated. NA (100 mu M) or methoxamine (100 mu M) evoked repeatable contraction s. Clonidine (100-300 mu M) was ineffective. The contractions to NA were re duced by procaine (1-10 mu M) but not by thapsigargin (0.1-0.3 mu M), ryano dine (1-30 mu M) or TMB-8 (1-30 mu M). Contractions to cumulative additions of NA (1-100 mu M) were enhanced in the presence of cyclopiazonic acid (10 & 30 mu M) but not ryanodine (10 & 30 mu M). Sequential contractions to NA were not blocked by PKC inhibitors, calphosin C (1 mu M) or Ro 31-8220 (1- 30 mu M) but were reduced by H-7 (1-30 mu M), a broad spectrum protein kina se inhibitor. Although RT-PCR experiments detected mRNA for some CA(2+)-dep endent/DAG-activated PKC isoforms in epididymal vas deferens, the PKC activ ators, phorbol 12, 13-dibutyrate (100 mu M) or phorbol 12-myristate 13-acet ate (100 mu M) failed to activate the tissues in calcium-free medium but en hanced subsequent contractions to NA. These results indicate a limited role for intracellular calcium stores and phorbol ester/DAG-sensitive PKC isofo rms in NA-induced contraction of epididymal rat vas deferens in calcium-fre e medium. The results suggest that pharmacomechanical coupling triggered by NA and CA(2+)-independent/DAG-activated may involve the sensitization of c ontractile myofilaments to Ca2+ or a Ca2+-independent mechanism. The possib le involvement of Ca2+-independent/DAG-insensitive PKC isoforms and agonist -dependent but PKC-independent sensitization pathway is discussed.