Abstract
Spontaneous purinergic neurotransmission was characterized in the mouse urinary bladder, a model for the pathological or ageing human bladder. Intracellular electrophysiological recording from smooth muscle cells of the detrusor muscle revealed spontaneous depolarizations, distinguishable from spontaneous action potentials (sAPs) by their amplitude (< 40 mV) and insensitivity to the L-type Ca2+ channel blocker nifedipine (1 μm) (100 ± 29%). Spontaneous depolarizations were abolished by the P2X1 receptor antagonist NF449 (10 μm) (frequency 8.5 ± 8.5% of controls), insensitive to the muscarinic acetylcholine receptor antagonist atropine (1 μm) (103.4 ± 3.0%), and became more frequent in latrotoxin (LTX; 1 nm) (438 ± 95%), suggesting that they are spontaneous excitatory junction potentials (sEJPs). Such sEJPs were correlated, in amplitude and timing, with focal Ca2+ transients in smooth muscle cells (measured using confocal microscopy), suggesting a common origin: ATP binding to P2X1 receptors. sAPs were abolished by NF449, insensitive to atropine (126 ± 39%) and increased in frequency by LTX (930 ± 450%) suggesting a neurogenic, purinergic origin, in common with sEJPs. By comparing the kinetics of sAPs and sEJPs, we demonstrated that sAPs occur when sufficient cation influx through P2X1 receptors triggers L-type Ca2+ channels; the first peak of the differentiated rising phase of depolarizations – attributed to the influx of cations through the P2X1 receptor – is of larger amplitude for sAPs (2248 mV s−1) than sEJPs (439 mV s−1). Surprisingly, sAPs in the mouse urinary bladder, unlike those from other species, are triggered by stochastic ATP release from parasympathetic nerve terminals rather than being myogenic.
Highlights
In most mammals, excitation of the detrusor muscle of the urinary bladder is achieved through the parasympathetic activation of both purinergic and cholinergic neurotransmission, leading to voiding of the bladder
Intracellular recording of smooth muscle cells in the mouse urinary bladder revealed both spontaneous depolarizations and spontaneous action potentials, as previously described. sAPs can be distinguished from sDeps by their greater amplitude, the presence of an after-hyperpolarization or after-depolarization and their sensitivity to the voltage-dependent Ca2+ channel (VDCC) antagonist nifedipine (Meng et al 2008)
In order to determine the mechanisms underlying the generation of sDeps, the contributions of purinergic and cholinergic neurotransmission were explored through the application of specific antagonists
Summary
Excitation of the detrusor muscle of the urinary bladder is achieved through the parasympathetic activation of both purinergic and cholinergic neurotransmission, leading to voiding of the bladder. The wide variation in reported contributions of purinergic neurotransmission to bladder voiding is partly explained by the methodological difficulty of evoking neural stimulation without evoking potentially masking direct muscle stimulation (Hoyle et al 1989). It is, generally agreed that purinergic neurotransmission is up-regulated in bladder dysfunction; Sjorgen et al (1982) demonstrated, for example, that while atropine abolished contractions in preparations from normal bladders, preparations from pathological bladders exhibited contractions that were up to 50% atropine resistant. It has been demonstrated that ATP, released from parasympathetic nerve terminals, binds to P2X receptors (Burnstock & Kennedy, 1985; Howson et al 1988) on urinary bladder smooth muscle cells (SMCs), producing an excitatory junction potential (EJP) that triggers the opening of L-type Ca2+ channels and the subsequent initiation of an action potential (Hoyle & Burnstock, 1985; Howson et al 1988). Heppner et al (2005) recently identified purinergic focal Ca2+ transients, corresponding to the influx of Ca2+ through P2X receptors upon ATP binding in rat urinary bladder smooth muscle
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