MLCK and PKC Involvements via Gi and Rho A Protein in Contraction by the Electrical Field Stimulation in Feline Esophageal Smooth Muscle

Abstract

W e have shown that myosin light chain kinase (MLCK) was required for the off-contraction in response to the electrical field stimulation (EFS) of feline esophageal smooth muscle. In this study, we investigated whether protein kinase C (PKC) may require the on-contraction in response to EFS using feline esophageal smooth muscle. The contractions were recorded using an isometric force transducer. On-contraction occurred in the presence of N G -nitro-L-arginine methyl ester (L-NAME), suggesting that nitric oxide acts as an inhibitory mediator in smooth muscle. The excitatory composition of both contractions was cholinergic dependent which was blocked by tetrodotoxin or atropine. The on-contraction was abolished in Ca 2＋ -free buffer but reappeared in normal Ca 2＋ -containing buffer indicating that the contraction was Ca 2＋ dependent. 4-aminopyridine (4-AP), voltage-dependent K＋ channel blocker, significantly enhanced on-contraction. Aluminum fluoride (a G-protein activator) increased on-contraction. Pertussis toxin (a Gi inactivator) and C3 exoenzyme (a rhoA inactivator) significantly decreased on-contraction suggesting that Gi or rhoA protein may be related with Ca 2＋ and K ＋ channel. ML-9, a MLCK inhibitor, significantly inhibited on-contraction, and chelerythrine (PKC inhibitor) affected on the contraction. These results suggest that endogenous cholinergic contractions activated directly by low-frequency EFS may be mediated by Ca 2＋ , and G proteins, such as Gi and rhoA, which resulted in the activation of MLCK, and PKC to produce the contraction in feline distal esophageal smooth muscle.

Lysophosphatidic acid presynaptically blocks NO uptake during electric field stimulation-induced relaxation via LPA1 receptor in cat lower esophageal sphincter

“…The present study shows that LPA is closely involved with endogenous NO release, which causes relaxation via an LPA 1 receptor-mediated pathway in LES, which is consistent with the findings of a previous study, in which it was found that EFS stimulates NO release (Park et al, 2009(Park et al, , 2010. Because recent studies have concluded that NO, rather than VIP, is the major mediator of nerve-induced smooth muscle relaxation (Uc et al, 1999), we examined the effects of LPA on the direct NO-induced relaxation of LES using SNP (a NO donor) as relaxant, and significant inhibition of SNP-induced relaxation was observed in an LPA concentration-dependent manner in cat LES.…”

Role of PKC and RhoA/ROCK pathways in the spontaneous phasic activity in the rectal smooth muscle

“…In contrast to the basal tone in human and cat lower esophageal sphincter, ACh-and EFS-induced phasic contraction in the esophageal body is primarily mediated by PKC (12,25,37). Similar conclusions were drawn by the inhibition of carbachol-induced contraction of urinary bladder by PKC inhibitor (7).…”

“…[In contrast to the phasic activity in RSM, a number of studies have shown that basal tone in the lower esophageal sphincter and IAS in humans and animals is mediated primarily via the RhoA/ROCK pathway (25,30,32,37).] The present data also show selectivity of the effects of PKC and ROCK inhibitors on the respective pathways.…”

P2X and P2Y Receptors Mediate Contraction Induced by Electrical Field Stimulation in Feline Esophageal Smooth Muscle

“…In vitro studies of human esophageal strips using direct activation of enteric motor neurons by electrical field stimulation (EFS) have shown that lower esophageal smooth muscle relaxation is mainly caused by NO released from inhibitory enteric motor neurons, whereas esophageal contraction is mainly caused by acetylcholine released from excitatory enteric motor neurons acting on muscarinic receptors located in the smooth muscle [3,[28][29][30]. Except for acetylcholine, other neurotransmitters influence EFS-induced contraction in esophageal smooth muscle.…”