Evidence against differential release of noradrenaline, neuropeptide Y, and dopamine‐β‐hydroxylase from adrenergic nerves in the isolated perfused sheep spleen

The subcellular storage and release of noradrenaline (NA), dopamine-beta-hydroxylase (D beta H), and neuropeptide Y (NPY) was studied in the isolated perfused sheep spleen. Subcellular distribution studies showed a bimodal distribution for NA which was well reflected by D beta H and indicated the occurrence of two types of NA storage vesicles. The most dense, presumably large dense-cored vesicles (LDV), contain both membrane-bound and soluble D beta H; the less dense presumably corresponds to small dense-cored vesicles (SDV) and at least does not contain soluble D beta H. The distribution of NPY is extended but shows a peak only at the position of LDV, indicating that LDV contain NPY. Continuous electrical stimulation of the splenic nerve at 2 Hz, 5 Hz, 10 Hz, and 20 Hz or at 20 hz with bursts induced the release of NA, NPY, and D beta H. The ratio among these components was constant. The fractional release of D beta H and NA was comparable at all frequencies used; that of NPY was 10-20 times lower, suggesting the occurrence of a large nonreleasable NPY pool. The present data argue against a high frequency stimulation or intermittent stimulation-induced preferential release of NPY from adrenergic neurons and question the concept of frequency-dependent chemical coding of sympathetic transmission in general. The simplest interpretation of our data is that NA and NPY are released at all frequencies from a single pool. The present finding might signify that only large dense-cored vesicles are involved in the sympathetic stimulation-evoked secretion of catecholamines from adrenergic nerve terminals of the isolated sheep spleen.