ABSTRACT
BACKGROUND AND PURPOSE: There is little information about the excitatory cholinergic mechanisms of mouse small intestine although this model is important for gene knock-out studies. EXPERIMENTAL APPROACH: Using patch-clamp techniques, voltage-dependent and pharmacological properties of carbachol- or intracellular GTPgammaS-activated cationic channels in mouse ileal myocytes were investigated. KEY RESULTS: Three types of cation channels were identified in outside-out patches (17, 70 and 140 pS). The voltage-dependent behaviour of the 70 pS channel, which was also the most abundantly expressed channel (approximately 0.35 micro(-2)) was most consistent with the properties of the whole-cell muscarinic current (half-maximal activation at -72.3+/-9.3 mV, slope of -9.1+/-7.4 mV and mean open probability of 0.16+/-0.01 at -40 mV; at near maximal activation by 50 microM carbachol). Both channel conductance and open probability depended on the permeant cation in the order: Cs+ (70 pS) >Rb+ (66pS) >Na+ (47 pS) >Li+ (30 pS). External application of divalent cations, quinine, SK&F 96365 or La3+ strongly inhibited the whole-cell current. At the single channel level the nature of the inhibitory effects appeared to be very different. Either reduction of the open probability (quinine and to some extent SK&F 96365 and La3+) or of unitary current amplitude (Ca2+, Mg2+, SK&F 96365, La3+) was observed implying significant differences in the dissociation rates of the blockers. CONCLUSIONS AND IMPLICATIONS: The muscarinic cation current of murine small intestine is very similar to that in guinea-pig myocytes and murine genetic manipulation should yield important information about muscarinic receptor transduction mechanisms.
Subject(s)
Ileum/metabolism , Myocytes, Smooth Muscle/metabolism , Receptors, Muscarinic/physiology , Transient Receptor Potential Channels/metabolism , Animals , Carbachol/pharmacology , Cholinergic Agonists/pharmacology , Female , Guanosine 5'-O-(3-Thiotriphosphate)/physiology , Ileum/cytology , Male , Mice , Mice, Knockout , Myocytes, Smooth Muscle/drug effects , Transient Receptor Potential Channels/geneticsABSTRACT
The carbachol-evoked inward cationic current in guinea-pig ileum smooth muscle cells is comprised of three types nonselective cationic channels (NSCC) with small (10 +/- 2 pS), medium (56 +/- 8pS) and large (135 +/- 14 pS) unitary conductance. All three types of NSCC could be activated by external application of carbachol as well as by internal application of GTPgS. It was found that behavior of carbachol- and GTPgammaS-evoked whole-cell current is mainly determined by the properties of medium conductance channels. The U-shaped I-V relationship of the whole-cell cationic current at negative potentials range arrives from voltage-dependence of its Po of this channel.
