Workshop on extrahypothalamic and hypophyseal neuropeptide sites and pathways.

The introduction by Coons in 1941 (7) of antibodies covalently coupled to fluorochromes as tracers for light microscopy initiated a period in which selective and specific staining of tissue compounds became possible. In spite of the fact that vasopressin (AVP) and oxytocin (OXT) had already been synthesized in 1953 (9), it was not until the beginning of the seventies that the first reports appeared regarding the immunocytochemical (ICC) localization ofpeptides in the nervous system (for references, see ref. 16). Before the introduction of ICC, the neurohypophyseal hormones (AVP, OXT, and their neurophysins) could be demonstrated with the method developed by Gomori ( 10). In this way Barry (3, 4) described in extensive comparative studies “Gomori-positive extrahypothalamic pathways” that were ending in various brain areas by “de synapses neurosecretoires. “ Although at that time, the (immuno)electron microscopic proof of such terminations did not exist, this important work has certainly not been adequately recognized. Hypothalamic releasing and inhibiting factors (e.g. , thyrotropinreleasing factor (TRF), luteinizing hormone-releasing factor (LRF), and somatotropin release inhibiting factor (SRIF)) were impossible to demonstrate by the conventional staining techniques. The elucidation of the chemical structure in the early seventies by Guillemin and Schally enabled the production of antisera, following the conjugation of the hormone to albumin or thyroglobulin (15). The same pattern holds for the pituitary hormones (e.g., ACTH/LPH family), enkephalins and peptides, that were not primarily known as hypothalamic or hypophyseal hormones (e.g., substance P. neurotensin, angiotensin II, vasoactive intestinal polypeptide, gastrin and cholecystokinin. In this way the previously described sites of synthesis of neuropeptides were confirmed and many new sites were discovered, resulting in a number ofnew concepts. It thus appeared that the synthesis and release of neuropeptides (releasing factors) was not restricted to a small population ofcells within the hypothalamus, the “hypophysiotropic area” (9), and the neurohypophysis, respectively, but is widespread throughout the brain, the spinal cord, and the peripheral nervous system. Peptides, formerly known to be present only in the adenohypophysis ( ACTH/LPH family, prolactin, growth hormone), are not only found to be present, but most probably also synthesized within the brain, as was already indicated by Zondek for intermedin in 1935 (21) (for references, see ref. 18), as are “tissue hormones” (e.g. , gastrin/cholecystokinin, angiotensin II) and “peripheral” nervous principles (substance P. vasoactive intestinal polypeptide) (5). Thus the quantity of peptides in the central nervous system appeared to outnumber the more classical transmitters, viz. the amines and amino acids, indicating extensive central functions for neuropeptides ( 12, 13). ICC suggested that the neuropeptides reached their central site of action not so much via the bloodstream or the cerebrospinal fluid, but rather directly via peptidergic pathways and synapses (Buijs, this issue, p. 357), Since the same neuropeptide can be localized in the brain at synaptic sites (Buijs, this issue, p. 357) and around capillaries in neurohemal regions (median cminence, neural lobe), the classifications of messenger substances into neurohumors and neurohormones ( 14) became outdated. From the field of neurophysiology, neuromodulators were introduced as a new type of messengers (e.g., ref. 2). In contrast to neurotransmitters, they do not act transsynaptically, need not have specific receptors, and are not responsible for direct transfer of action potentials, but affect such actions as neurotransmitter synthesis and release instead ( 1, 6). It appeared, however, from ICC observations, that neuropeptides (e.g., enkephalins, AVP and OXT) were found in synapses that were not distinguishable from classical aminergic synapses, suggesting that they may act as neurotransmitters as well. In the past years an enormous number of publications on ICC of neuropeptides have appeared. There have been a number of contradictory reports (e.g., the presence ofsomatostatin (SOM or SRIF) in magnocellular cells of the paraventricular and supraoptic nucleus (8)). It seems therefore appropriate to single out problems of serum specificity as a possible source for discrepancies in the literature. In most papers, as proof of serum specificity (viz. th exclusive immunological binding to the compound to which the antibody was raised), the antiserum is only absorbed to the homologous antigen. This absorption test, however, proves nothing more than that all antibodies were capable of