Bretylium or 6‐OHDA‐resistant, action potential‐evoked Ca²⁺ transients in varicosities of the mouse vas deferens

Abstract

Action potential-evoked calcium transients in varicosities in mouse vas deferens were monitored using laser scanning confocal microscopy. Their signi®cance was examined by comparison with excitatory junction potentials (EJPs) and neurogenic contractions, both indirect measurements of transmitter release. Bretylium abolished EJPs, as well as the ATP and NA-mediated phases of contraction. However, bretylium revealed a prominent late component of contraction that was atropine-sensitive. Bretylium abolished calcium transients in 21%, enhanced in 16% and had no e ect in 63% of varicosities examined. Pre-treatment with 6-OHDA reduced NA levels to below detectable levels but many strings of varicosities still responded to nerve impulses with`normal' calcium transients. Varicosities in which calcium transients were abolished by these agents were sympathetic. The identity of those varicosities in which calcium transients were resistant to bretylium (sympathetic but no uptake-1 sites, parasympathetic, sensory) remains to be established.

Cholinergic innervation of the guinea-pig isolated vas deferens

“…Bretylium greatly reduced both the purinergic and noradrenergic components of electrically evoked responses. Continuous exposure (approximately 2–3 h) to bretylium revealed a residual hump-like contraction (at 200 stimuli), which increased in amplitude with time, reaching a maximum after about 5 h (see Jackson and Cunnane 2002) (Fig. 1, n =12, P <0.05).…”

“…In mouse vas deferens, contraction and confocal studies have shown that nicotine increases the amplitude of the bretylium-resistant contraction (Cuprian et al 2005) and spontaneous Ca 2+ transients in all varicosities, located on either bretylium-sensitive or bretylium-resistant nerve terminals (Brain et al 2001; Jackson et al 2001; Jackson and Cunnane 2002). These nerve terminals were considered to be preterminal sympathetic fibres, sensory nerves or cholinergic nerves.…”

Peripheral Nervous System

“…Nerve fibres, originating from pelvic neurons in a discreet location in the pelvic ganglia supply the rat vas deferens (Keast 1992). A population of varicose nerves has been identified in the mouse vas deferens after destruction of sympathetic nerves by 6-hydroxydopamine (6-OHDA) (Jackson and Cunnane 2002). These nerves may be parasympathetic and/or sensory afferent fibres arising from L1, L2, L6 and S1 DRG travelling to the rat vas deferens in both hypogastric and pelvic nerves.…”

Bretylium abolishes neurotransmitter release without necessarily abolishing the nerve terminal action potential in sympathetic terminals

“…It has been argued that bretylium ( Boura and Green, 1959 ) prevents neurotransmitter release from sympathetic nerve terminals by abolishing the nerve terminal action potential ( Haeusler et al , 1969 ; Mitchell and Oates, 1970 ; Maxwell and Wastila, 1977 ; Brock and Cunnane, 1988 ); selectivity for sympathetic terminals occurs because bretylium is taken up into sympathetic terminals by a norepinephrine (NA) transporter‐1 (or uptake‐1). Hence, a potentially useful approach to identify noradrenergic terminals is to detect strings of varicosities in which the nerve action potential is abolished by bretylium ( Jackson and Cunnane, 2002 ). However, Jackson and Cunnane (2002) showed that bretylium (20 μ M ) does not abolish the action potential‐induced Ca 2+ transient in the majority of nerve terminal varicosities orthogradely loaded with the dextran conjugate of a Ca 2+ indicator.…”

“…Hence, a potentially useful approach to identify noradrenergic terminals is to detect strings of varicosities in which the nerve action potential is abolished by bretylium ( Jackson and Cunnane, 2002 ). However, Jackson and Cunnane (2002) showed that bretylium (20 μ M ) does not abolish the action potential‐induced Ca 2+ transient in the majority of nerve terminal varicosities orthogradely loaded with the dextran conjugate of a Ca 2+ indicator. This causes a dilemma because the innervation of the mouse vas deferens is predominantly sympathetic ( Taxi, 1965 ; Jones and Spriggs, 1975 ).…”