Calcium in sympathetic varicosities of mouse vas deferens during facilitation, augmentation and autoinhibition.

Abstract

1. The sympathetic nerve terminals of the mouse vas deferens were loaded with the calcium indicator Oregon Green 488 BAPTA-1 by orthograde transport along the postganglionic nerves. Changes in the calcium concentration in the varicosity (delta [Ca2+]v) were determined following single impulses, and short (5-impulse) and long (200-impulse) trains at 5 Hz. 2. All varicosities showed a significant delta [Ca2+]v in response to every single impulse. The elevated delta [Ca2+]v declined in two phases with similar kinetics for all varicosities: a fast phase (time constant, 0.42 +/- 0.05 s) and a moderate phase (3.6 +/- 0.4 s). 3. Line scanning confocal microscopy revealed that the delta [Ca2+] of a single terminal following single impulses was smaller for the intervaricose regions than for the varicosities. 4. Blockade of the voltage-sensitive calcium channels with Cd2+ (in calcium-free solution) completely blocked the delta [Ca2+]v on stimulation. The addition of either nifedipine (10 microM), omega-conotoxin GVIA (100 nM) or omega-agatoxin TK (100 nM) showed that 47 +/- 6% of the evoked response was mediated by N-type calcium channels. 5. Ryanodine (10 microM) did not significantly change the amplitude of delta [Ca2+]v in response to short trains. 6. Spontaneous increases in delta [Ca2+]v were observed in individual varicosities, with coupling in the increase of delta [Ca2+]v between varicosities. 7. The presynaptic alpha 2-receptor antagonist yohimbine (10 microM) increased the amplitude of delta [Ca2+]v in response to five impulses (5 Hz) by 54 +/- 14%, while the alpha 2-receptor agonist clonidine (1 microM) decreased the delta [Ca2+]v by 55 +/- 4%. 8. These results are discussed in terms of the hypotheses that the increased probability for secretion at sympathetic nerve terminals which accompanies facilitation and augmentation is due to the residual delta [Ca2+]v remaining after the calcium influx following impulses and that noradrenaline acts presynaptically to decrease the probability of secretion by modifying calcium influx.