Pharmacological evidence that GABA-induced relaxation of rat proximal duodenum longitudinal muscle depends on NKCC cotransporter activity and Ca2+ influx

Abstract

γ-Aminobutyric acid (GABA) is the main inhibitory neurotransmitter in adult central nervous system (CNS) synapses, but it excites immature CNS neurons as well as neurons in the myenteric plexus. The present work aimed to determine whether GABA-induced nonadrenergic, noncholinergic (NANC) neuronal-mediated relaxation of the rat duodenum is dependent on the activity of Na+ K+ Cl- cotransporters (NKCC) and requires calcium influx. In the presence of guanethidine (3 µmol/L), atropine (3 µmol/L), and indomethacin (1 µmol/L), relaxations induced by GABA (100 µmol/L), KCl (5–10 mmol/L) and electrical field stimulation (1–8 Hz, 2 ms, 60 V), but not those induced by bradykinin (10–100 nmol/L) were abolished by lidocaine (300 µmol/L). However, only GABA-induced relaxations were reduced in a concentration-dependent manner by the NKCC1/2 inhibitors bumetanide (0.1–1 µmol/L) and furosemide (1–10 µmol/L). GABA-induced NANC neuronal relaxation was abolished by bicuculline (30 µmol/L) and inhibited by N-nitroarginine methyl ester (l-NAME, 300 µmol/L). The ω-conotoxin GVIA (1 µmol/L), which acts exclusively on neuronal CaV2 channels, but not on smooth muscle voltage-gated Ca2+ CaV1 channels, and nonselective blockers of these channels (verapamil 100 nmol/L and ruthenium red 10 µmol/L), reduced GABA-induced relaxations. These results showed that the activation of GABAA receptors induces NANC nitrergic neuronal relaxations in the rat duodenum, which depend on NKCC activity and CaV2 channel activation, suggesting that this phenomenon results from neuronal depolarization promoted by Cl− efflux through GABAA receptors, with subsequent Ca2+ influx and nitric oxide release.