The overall goal of this proposed research is to contribute to basic biochemical understanding of the pharmacological activity and toxicology of marijuana and its major phychoactive component, delta9-tetrahydrocannabinol (delta9-TCH). This should contribute to better understanding of toxic effects of marijuana and of methods of preventing marijuana abuse and possibly to the development of therapeutic drugs with the benefits of tetrahydrocannabinol but without the undesirable side effects. It is proposed to achieve these objectives by means of a systematic search for cannabinoid receptors which are hypothesized to mediate some of the drug's pharmacological actions. Antibodies to delta 9-THC will be used to immunize animals. Based on published precedents, it is hypothesized that this will stimulate the formation of antibodies, some of which will contain within their structure a reflection of the binding site of the primary antibody. This substructure may be regarded as an internal peptide image of delta9-THC and it will bind to the cannabinoid binding site of the original antisera. Appropriately structured antibodies with an internal THC image should also bind to the hypothesized THC receptors. By use of hybridoma techniques, a panel of monoclonal antibodies of this type will be developed. Techniques such as radioisotope, fluorescent, enzyme or colloidal gold labeling of these antibodies, as well as second antibody techniques for localization will be developed. The interactions of these THC internal image antibodies with tissues, cells and subcellular fractions will then be studied in an attempt to find receptor sites for cannabinoids. The specificity of these receptors will be tested by studying the inhibition of the binding by cannabinoids and other pharmacologically active molecules. Binding will also be studied after in vivo administration of the antibodies, mainly by intracerebroventricular techniques. Electron microscopy will aslo be used to help define localization of the receptor sites. Locomotor activity and hypothermia will be used as tests for in vivo agonist or antagonist activity of the antibodies versus tetraphdrocannabinol and discriminative stimulus poperties will be tested.
