Vasostatins, N‐terminal products of chromogranin A, are released from the stimulated calf spleen in vitro

Vasostatins are the N‐terminal chromogranin A peptides 7 ˜ 22 kDa. They have been shown to be present in several endocrine tissues and exhibit vasoinhibitory activity in vitro. In a first series of experiments, we investigated the presence and subcellular localization of vasostatins in the bovine splenic nerve. Experimental results, obtained using gradient centrifugation, showed that noradrenaline was enriched 25‐fold in the large dense core vesicle fraction, compared with the original homogenate. In the latter fraction, the 7 and 18 kDa peptides were observed following immunodetection with antiserum to chromogranin A_{1, 4o} and laser densitometric scanning revealed these two fragments as the major N‐terminal fragments. Subsequently, we examined the release of the 7 and 18 kDa peptides from perfused calf spleen during veratridine (20 μM) or 1,1‐dimethyl‐4‐phenylpiperazinium iodide (20 μM) stimulation. In the prestimulatio samples, we were not able to detect these peptides, however, following stimulation, the 7 and 18 kDa chromogranin A fragments became apparent. The vasostatin‐immunoreactivity, in both bovine chromaffin granule lysate and calf spleen perfusate, elutes at the same retention time on reversed‐phase high performance liquid chromatography. The present study demonstrated that vasostatins are present in the large dense core vesicles of sympathetic axons and are released from the nerve terminals in response to stimulation. The release of vasostatins from sympathetic nerves in the spleen suggest an in vivo function for N‐terminal chromogranin A products of neuronal origin.