Ultrastructural localization of enkephalin‐like immunoreactivity in the rat adrenal medulla

Abstract

The PAP method was applied to adrenomedullary cells to demonstrate the subcellular localization of ELI. The labeled cell-groups were identified from flat-embedded virbratome sections by light microscopy. Electron microscopy showed that the precipitate was associated with the storage granules of the cells. The number of labeled granules varied greatly from cell to cell. In cells corresponding to those showing heavy staining by light microscopy, all the granules were stained with precipitate. It is suggested that the peptide responsible for the ELI might be released together with the catecholamines.The pentapeptides (Met') and (Leu') enkephalins have been found widely distributed in the peripheral autonomic nervous system, as well as in the central nervous system (Schultzberg et al., '79; Miller and Cuatrecases, '79). Adrenomedullary cells of the rat and guinea pig The subcellular localization of ELI is described in this paper.MATERIALS AND METHODS The adrenals of white male rats of Sprague-Dawley strain were removed under light ether anesthesia, sliced, and immersed in phosphate-buffered (0.1 M, pH 7.3) 4% formaldehyde with 0.2% glutaraldehyde at 4°C. The fixative was freshly made from paraformaldehyde powder and highly purified glutaraldehyde. After fixation (60-120 min) and washing (24 h) the tissue was sectioned on a vibrating microtome (VIBRATOME@ in the buffer solution at 4°C. The 20-to 50-pm slices were processed through immunohistochem cal staining according to a modification (Pickel et al., '75, '79) of the peroxidase-antiperoxidase (PAP) method of Sternberger ('74). Controls included incubations with normal rabbit serum and antigen-inactivated serum (50 pg of leu-enkephalin in 1 ml of diluted antiserum).plastic coverslips in Epon. After the light-microscopic (LM) identification of labeled cells (Figs. 1, 2), the selected areas of the vibratome section were cut under a dissecting microscope and reembedded for ultramicrotomy. No grid staining was used.The antiserum against (Leu') enkephalin was a gift from Dr. Richard J. Miller, University of Chicago, and a detailed description of the specificity of the antiserum is given in Miller et al. ('78a, b).The sections were postfixed with 2% Os04 for 1 hour and embedded flat between two

Immunochistochemical Analysis of the Localization of Neuropeptides in the Adrenal Gland

“…Leu-ENK-like immunoreactivity has been shown in the in-situ chromaffin cells of rats, guinea pigs, hamsters, cats, bovines and men (SCHULTZBERG et al, 1978;HERVONEN et al, 1980;LINNOILA et al, 1980;LIVETT et al, 1982;PELTO-HUIKKO et al, 1982). Met-and Leu-ENK-like immunoreactivities have also been detected in the cultured chromaffin cells of bovines (LIVETT and DEAN, 1980;LIVETT et al, 1981;SIEGEL et al, 1985).…”

Effect of Endogenous Opioid Blockade on Fetal Cardiovascular and Sympathoadrenal Responses to Hypoxemia Induced by Umbilical Cord Constriction

“…Plasma catechol amines rose less in group IV than in hypoxemic control fetuses (norepinephrine: 2.407 ± 406 vs. 3,410 ± 617 pg/ml; epinephrine: 966 ± 641 vs. 1,074 ± 305 pg/ml), but the differences were not significant (p > 0.2). It is concluded that endogenous opioids do not play a major role in the modulation of cardiovascular and sympathoadrenal adaptions to moderate hypox emia in the fetus.The cardiovascular responses of the fetal sheep to hypoxemia are mediated by pro found increases in parasympathetic and sym pathoadrenal activity [1,2], Endogenous opioids and their receptors are distributed widely in the brain and the adrenal medulla, and several investigators have suggested that endorphins and enkephalins may act as ncuromodulators of cathecholamine release [3][4][5][6][7], In adult animals, opioid receptor block ade improves circulation impaired by the se- …”

Catecholamine and Endorphin Responses to Delivery in Term and Preterm Lambs

“…Within the central nervous system, en kephalin and endorphin containing neurons are found in hypothalamic nuclei impor tantly involved in autonomic regulation [26], Endogenous opiates inhibit neurotrans mission in sympathetic ganglia [14,15], They also inhibit adrenal medullary cate cholamine release with ß-END being several orders of magnitude more effective than morphine and effective at doses comparable to those seen during a variety of altered physiologic states [16]. Finally, there is evi dence that endorphin secretion exacerbates the circulatory collapse in a number of shock models and that naloxone can reverse the collapse [25][26][27][28][29], Moreover, an intact sym pathoadrenal system is necessary for the ameliorative effects of naloxone [30].…”

“…Opiate peptides, largely metenkephalin, are localized in the peripheral nervous system in sympathetic ganglia [22,23], preganglionic neurons [24] and the adrenal medulla [23][24][25]. Within the central nervous system, en kephalin and endorphin containing neurons are found in hypothalamic nuclei impor tantly involved in autonomic regulation [26], Endogenous opiates inhibit neurotrans mission in sympathetic ganglia [14,15], They also inhibit adrenal medullary cate cholamine release with ß-END being several orders of magnitude more effective than morphine and effective at doses comparable to those seen during a variety of altered physiologic states [16].…”