Innervation of the sheep pineal gland by nonsympathetic nerve fibers containing NADPH-diaphorase activity

We used the NADPH-diaphorase histochemical method as a potential marker for nitric oxide synthase (NOS)-containing nerve fibers innervating the pineal gland of the sheep. Nerve fibers containing NADPH-diaphorase activity provide dense innervation of the sheep pineal gland. The nerve fibers were located in the pineal capsule, in the connective tissue septae separating the lobuli of the gland, and penetrating between the pinealocytes. The nerve fibers were either smooth or endowed with boutons en passant. After bilateral removal of the superior cervical ganglion, the dense network of NADPH-diaphorase-positive fibers was still present in the gland. Ganglionectomy affected neither the distribution nor the appearance of the NADPH-diaphorase-positive fibers. Most of the NADPH-diaphorase-positive fibers also contained peptide histidine isoleucine and vasoactive intestinal polypeptide, and a comparatively smaller fraction contained neuropeptide Y. Pinealocytes never exhibited NADPH-diaphorase activity. These results demonstrate a major neural input to the sheep pineal gland with NADPH-diaphorase-positive nerve fibers of nonsympathetic origin.