Maxadilan is the most potent vasodilator known. It is a novel peptide recently isolated from salivary glands of the sand fly Lutzomyia longipalpis, a vector of the protozoan disease leishmaniasis. In this proposal, studies are directed at finding the receptor for maxadilan, examining the pharmacologic actions of this peptide, and characterizing its structural and functional domains. The receptor for maxadilan appears to be on vascular smooth muscle cells and preliminary studies suggest that maxadilan raises the level of cAMP in these cells. Elevation in the intracellular level of this second messenger probably results from the activation of G-protein coupled receptor by maxadilan. This receptor may be important in the physiological regulation of blood vessel tone but is unlikely to have been described previously and the endogenous ligand for this receptor is not known. Expression of cDNA libraries derived from rabbit aorta or a cell line to which maxadilan binds, will be performed in COS cells in order to obtain the gene encoding the maxadilan receptor. Receptor distribution will be determined by probing northern blots from various tissues with the receptor probe while autoradiography of skin probed with radiolabelled maxadilan will allow for microlocalization of the receptor. Pharmacology studies are designed to examine the effect of maxadilan on classical vascular smooth muscle preparations and define the second messenger pathway stimulated by this peptide. The various pharmacologic agents including channel blockers and receptor blockers. The capacity of maxadilan to alter second messenger levels will be determined in selected vascular smooth muscle preparations or cultured cells by radioimmunoassay of cAMP and cGMP levels, calcium flux measurements, and the HPLC-based detection of radiolabeled inositol phosphates. Structural and functional domains of maxadilan will be studied by generating monoclonal antibodies to the peptide, expressing altered forms of the peptide with recombinant techniques and determining the structure of maxadilan by 2-dimensional nuclear magnetic resonance. The long-term objectives of this laboratory are to exploit the mechanisms by which arthropods successfully obtain blood. The overriding concept is that an organism (arthropod) can be examined for a particular function (blood-feeding) leading to discovery of novel proteins (maxadilan) and genes. Such proteins and genes will provide new tools with which to study hematologic and cardiovascular pharmacology and will lead to new therapeutic agents.
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