Gap junction blockade with carbenoxolone differentially affects fictive breathing in larval and adult bullfrogs.

Abstract

This study examined the role of gap junctional communication in the modulation of respiratory related motor output using in vitro brainstem preparations of larval (N=14) and adult (N=14) bullfrogs (Rana catesbeiana). Superfusion of the isolated brainstem for at least 1 h with the gap junction blocker carbenoxolone (CBX; 100 microM and 1 mM) dissolved in artificial cerebrospinal fluid (aCSF) elicited significant changes in respiratory-related burst frequency in both larval and adult preparations. In tadpole preparations, both concentrations of CBX significantly decreased gill and lung burst frequency over 20-40 min, with 1 mM CBX producing complete cessation of gill and lung burst activity by 40 min in all preparations. There was little or no change in other burst characteristics such as burst amplitude or duration. By contrast, superfusion of the adult brainstem preparation with CBX significantly increased lung burst frequency over 10-20 min, and caused cessation of lung burst activity with 100 microM CBX (five of seven preparations) and with 1 mM CBX (seven of seven preparations). Adult preparations that ceased activity with 100 microM CBX recovered in control aCSF, but those in 1 mM did not recover, despite up to 3 h superfusion with control aCSF. In two additional adult preparations, 1 h exposure to hypercapnic aCSF (7-10% CO2) following the cessation of fictive breathing with 1 mM CBX failed to evoke respiratory activity. The inhibition of fictive breathing in tadpoles suggests that gap junctional communication may be important for respiratory rhythmogenesis prior to the development of central CO2 chemosensitivity. Following metamorphosis to the terrestrial adult, however, gap junctional communication may contribute to regulation of respiratory frequency and possibly the transduction of central CO2 chemosensitivity.