Relatively little is known about the contribution of Ca²⁺ dependent and independent mechanisms in the contractility of neonatal smooth muscle. We have therefore studied Ca²⁺ homeostasis and Ca²⁺ sensitization mechanisms in 10-day and adult guinea pig gallbladder smooth muscle to elucidate developmental changes in these processes. Gallbladder contractility was evaluated by isometrical tension recordings from strips, [Ca²⁺ ]i was estimated by epifluorescence microscopy of fura-2 loaded isolated cells and protein expression and phosphorylation was assessed by western blot.

The neonatal gallbladder contracted significantly less to KCl and cholecystokinin (CCK) than adult tissue. KCl-induced impaired contractility correlated with a decrease in Ca²⁺ influx but, surprisingly, CCK-evoked response was associated with increased Ca²⁺ mobilization from intracellular stores, suggesting an impairment in Ca²⁺ sensitization mechanisms. The enhanced Ca²⁺ release in newborn gallbladder was the result of the increase in the size of the releasable Ca²⁺ pool, the decrease in the activity of extrusion mechanisms through plasma membrane and the over-activity of SERCA pump. The reduced Ca²⁺ sensitivity in neonatal tissues was demonstrated by the lack of effects Y27362, an inhibitor of Rho kinase (ROCK) and GF109203X, an inhibitor of protein kinase C (PKC), on agonist-induced contraction. In addition, neonatal gallbladder showed lower levels of RhoA, ROCK, PKC and the two effectors CPI-17 and MYPT1, and the absence of CPI-17 and MYPT1 phosphorylation in response to agonists.

In conclusion, our results indicate that the main mechanisms involved in smooth muscle contractility are under developmental regulation. Supported by BFU2004-0637.