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cultures. The structural basis for the helper signal may reside in the six N-terminal amino acids of AVP based on the relative abilities of AVP, oxytocin, vasotocin, and pressinoic acid (AVP six N-terminal amino acid peptide) to provide maximal help at 10 -9 to 10 -1° M. Antagonists of V 1 (hepatic) and V 2 (renal) AVP receptors were tested for their relative abilities to block the AVP helper signal for IFN-7 production. V 2 antagonists did not have an effect on AVP help, even at a 500-fold excess of antagonist. Two strong V 1 antagonists, [(1-fl-mercapto-fl,fl-cyclopentamethylene propionic acid)2-(O-methyl)tyro- sine,8-arginine] vasopressin ([d(CH2)aTyr(Me) 2] AVP and [1-(fl-mercapto-fl,fl-cyclopentamethylene propionic acid)2-D-(O-ethyl)tyrosine,4-valine,8-arginine] vasopressin ([d(CH2)D-Tyr(Et)2,Val 4] AVP), were also tested, but only [d (CH 2 )ITyr(Me) 2] AVP blocked AVP help. These data suggest that the AVP helper signal operates via a novel Vl-like AVP receptor on lymphocytes. Functional interaction of AVP with its lymphocyte receptor was specifically blocked by a nonapeptide that was 'coded' for by the AVP anti-sense D N A strand. This observation should provide a mechanism for isolation and characterization of the AVP receptor.

8. Does serotonin modulate macrophage function? Role of 5-HT binding sites and uptake mechanisms - - J.C. Jackson, W.H. Brooks and T.L. Roszman (University of Kentucky College of Medicine, Lexington, KY, U.S.A.)

Recently, we confirmed and extended observations that alterations in serotonin levels in vivo can influence immune function. Here, we report specific association of serotonin (5-HT) with lymphoid cells in the CBA mouse. Macrophages, but not T or B cells, appear to accumulate 5-HT via a high-affinity ( K m = 30 nM), active uptake mechanism. This system appears to be specific in that unlabelled 5-HT and the selective 5-HT uptake blocker, fluoxetine are most effective in competition studies. The 5-HT antagonists spiperone, ketanserin, cinanserin, and cyproheptadine also demonstrate significant capacity to inhibit uptake, but with lower affinity. Currently, evidence to support high-affinity 5-HT binding to lymphocyte membranes is lacking. Additionally, we have demonstra ted dose-dependent suppression by 5-HT of "r-INF-induced expression of Ia antigen in thioglycolate-elicited peritoneal macrophages, a phenomenon which can be prevented by both fluoxetine and 5-HT antagonists. Finally, HPLC studies have revealed that macrophages are able to metabolize 5-HT to the 5-hydroxyindoleacetic acid metabo- lite (5-HIAA). We therefore propose that macrophage function can be modulated through specific interaction with 5-HT, probably via one or more second messenger systems, and possibly incorporating metabolism of the amine. Serotonin may, in concert with other immune mediators, thereby influence immune processes.

9. Thymic hormone stimulation of pituitary hormone release - - B.L. Spangelo and R.M. MacLeod (Department of Medicine, University of Virginia School of Medicine, Charlottesville, VA 22908, U.S.A.)

Thymosin fraction 5 (TF5) is a partially purified extract of bovine thymus that contains 40-60 peptides. Beyond its well documented irmnunopotentiat ing effects, TF5 has recently been shown to affect the secretion of some hypothalamic peptides and pituitary hormones. TF5 (1-100 # g / m l ) stimulated prolactin and growth hormone (GH) release from both normal anterior pituitary and MtTW15 tumor cells, but had no effect upon luteinizing hormone release. TF5 st imulation of prolactin release from 45Ca2 + prelabeled perifused normal anterior pituitary cells was dose responsive and could be described as a 'block wave'; however, no concurrent change in 43 Ca 2 + efflux was observed. In contrast, the calcium channel blocker D-600 was found to inhibit TF5-mediated hormone release from normal anterior pituitary cells. We found that TRH-st imula ted prolactin release and GRF-s t imula ted GH release were further enhanced by TFS, suggesting a mechanism of action different from these peptides. Dopamine (500 nM) was found to block TF5-stimulated prolactin release from normal anterior pituitary cells, but SRIF (10-100 nM) had no effect on TF5-st imulated prolactin or G H release. TF5 failed to affect either basal or TRH-induced polyphosphoinosit ide hydrolysis. TF5 also stimulated the release of prolactin from 7315a tumor cells, but no changes in intracellular cAMP or cGMP levels could be correlated with these endocrine effects. BW755c, an inhibitor of all known catabolic pathways of arachidonic acid, was found to reduce the ability of TF5 to stimulate both prolactin and GH release. These data suggest that the

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stimulus secretion coupling mechanism does not involve polyphosphoinositide hydrolysis or cAMP generation, but appears to be dependent upon the generation of arachidonate metabolites.

10. Innervation of lymphoid tissue - - Suzanne Y. Felten and David L. Felten (University of Rochester School of Medicine, Department of Anatomy, Rochester, NY 14642, U.S.A)

Direct neuronal connections between the central nervous system and various primary and secondary lymphoid tissues have been demonstrated using a number of anatomical techniques used separately and in combination, including histochemistry, histofluorescence, immunocytochemistry, electron microscopy, immunoelectron microscopy, and pathway tracing. Studies of this nature have provided clear and unequivocal evidence that sympathetic postganglionic autonomic nerve fibers innervate thymus, spleen, and lymph nodes. These nerve fibers arise from sympathetic chain ganglion such as the superior cervical (thymus) or from collateral ganglia such as the coeliac-superior mesenteric (spleen), which, in turn receive input from the intermediolateral cells column of the thoracohimbar spinal cord. The intermediolateral cells column, in turn, receives projections from central autonomic sites such as the paraventricular nucleus of the hypothalamus, and the dorsal vagal complex. The noradrenergic sympathetic innervation enters the organ in association with blood vessels, but branches from these vessels to innervate the lymphatic parenchyma as well. It is most often closely associated with zones of lymphoid tissue containing T lymphocytes, including both the cortex and corticomedullary boundary of the thymus, the white pulp of the spleen, and the non-nodular regions of the lymph node. In the spleen, where the innervation has been investigated in great detail, the noradrenergic nerve fibers are specifically associated with the periarteriolar lymphatic sheath, the inner marginal zone, and the parafollicular zone in addition to innervation of the capsule, trabeculae and vasculature. Studies at the EM level confirm that noradrenergic fibers are in direct contact with lymphocytes. In the lymph node, capsular, paracortical zones, and medullary areas, particularly near the hilus are also innervated. The clear existence of a neuronal pathway by which the CNS might influence cells of the immune system provides a much needed additional link between psychosocial factors impacting on an individual and the outcome of disease. (Supported by N00014-84-K-0488 from the Office of Naval Research)

11. Nonadrenergic neurotransmission in the spleen - - David L. Felten and Suzanne Y. Felten (Depart- ment of anatomy, University of Rochester School of Medicine, Rochester, NY 14642, U.S.A.)

The noradrenergic innervation supplying the spleen fulfills the major criteria for neurotransmission, with lymphoid cells as the target tissue. Anatomically, noradrenergic fibers derived from coeliac-superior mesenteric ganglion cells, enter the spleen with the vasculature, and distribute into specific compartments of the white pulp. Double labelled immunocytochemistry has shown tyrosine hydroxylase-positive fibers adjacent to T lymphocytes in the periarteriolar lymphatic sheath, and adjacent to ED-3-positive macrophages at the marginal sinus. The spleen contains a high concentration of NE and its metabolites, a content that can be depleted by 6-hydroxydopamine, suggesting that it is found in the neural compart- ment. In vivo dialysis measurements of NE available in the extracellular space demonstrates the availability of a high concentration, in the range of 1 /~M, of NE, apparently derived from release. In addition lymphocytes possess beta-adrenergic receptors linked with cyclic AMP as a second messenger, and are able to alter intracellular metabolic events upon stimulation. At a functional level, in adult mice denervated with 6-hydroxydopamine, primary and secondary antibody responses are markedly di- minished in spleen and lymph nodes, suggesting that these nerves are important for full immunocompe- tence. In the aged Fischer 344 rat, the splenic noradrenergic innervation is reduced in all compartments, particularly in the parenchymal compartment, and the content of NE is reduced. The possibility of a causal relationship between diminished sympathetic noradrenergic innervation and diminished immune responsiveness in these aged rodents is currently being explored. We also have challenged lymphocytes in order to observe the potential plasticity for the NE fibers innervating the white pulp. If lymphocytes are destroyed with hydrocortisone or cyclophosphamide, the splenic weight diminishes and the white pulp shrinks, but the splenic NE content remains constant; however, the fiber density increases in the white pulp as the fibers retract to maintain their original compartmentation, indicating that replenishing lymphocyte populations will encounter higher concentrations of NE than in control spleens. If lympho-