Phorbol ester effects on coupling mechanisms during cholinergic contraction of swine tracheal smooth muscle.
1. We studied effects of the phorbol ester, phorbol 12,13-dibutyrate (PDB), on carbachol-induced contractions of swine trachealis muscle. PDB (1-10 microM) markedly inhibited 5.5 microM-carbachol-induced inositol phosphate synthesis allowing us to study (a) whether the membrane potential-independent component of force (pharmacomechanical coupling component) developed in carbachol-stimulated trachealis muscle is dependent on activation of inositol phospholipid metabolism, and (b) whether carbachol-induced membrane depolarization and contraction are altered in muscle where second messenger signals generated by inositol phospholipid metabolism are inhibited and activation of protein kinase C (PKC) is already maximal. 2. Application of PDB (10 microM) to unstimulated trachealis muscle resulted in a small slowly developing contraction associated with a 10 m V membrane depolarization. PDB-evoked contractions were not influenced by Na+ or Cl- ion substitutions, or administration of amiloride, all of which inhibited PDB-evoked membrane depolarization. 3. Pre-treatment with PDB had no effect on [K+]-force, or [K+]-membrane potential relationships, over a range of extracellular [K+] from 40 to 70 mM. Pretreatment with PDB had no effect on extracellular [Ca2+]-force relationships during 40 mM-K+. 4. Carbachol-evoked contractions of muscle treated with PDB became similar to K+ contractions in regard to effects of organic Ca2+ antagonist drugs or decrease in bathing solution [Ca2+]. At low carbachol concentrations, verapamil plus PDB completely inhibited force development. With 5.5 microM-carbachol, over 90% of total carbachol-induced force was inhibited by verapamil, or nifedipine, plus PDB. 5. Control carbachol-evoked contractions were associated with 20-25 mV membrane depolarizations. In PDB-treated muscle, carbachol-evoked contraction occurred with a blunted depolarization, i.e. about 5 mV. 6. Force controlled by pharmacomechanical coupling mechanisms operating during maintained carbachol-evoked contractions was inhibited by treatment with PDB. Carbachol-induced force dependent on pharmacomechanical coupling mechanisms could be explained by signals generated via inositol phospholipid metabolism. 7. Electromechanical coupling mechanisms were augmented during carbachol in PDB-treated muscle. This appears to be due primarily to changes in the properties or number of surface membrane voltage-gated Ca2+ channels. 8. Data suggest an important role of PKC-mediated phosphorylations for control of both pharmacomechanical coupling mechanisms mediated by activation of inositol phospholipid metabolism and electromechanical coupling mechanisms mediated by effects on operation of surface membrane ion channels.
Baba
K
K
Department of Physiology, School of Medicine, University of Pennsylvania, Philadelphia 19104.
Baron
C B
CB
Coburn
R F
RF
D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D002217:Carbachol / Q000494:pharmacology*; D066298:In Vitro Techniques; D007295:Inositol Phosphates / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D009119:Muscle Contraction / Q000187:drug effects*; D009130:Muscle, Smooth / Q000187:drug effects* / Q000378:metabolism / Q000502:physiology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology*; D013552:Swine; D014132:Trachea / Q000187:drug effects / Q000378:metabolism / Q000502:physiology; D014700:Verapamil / Q000494:pharmacology
D007295:Inositol Phosphates; D015240:Phorbol 12,13-Dibutyrate; D000584:Amiloride; D002217:Carbachol; D014700:Verapamil