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This paper describes the interaction of apamin, a bee venom neurotoxin, with the mouse neuroblastoma cell membrane. Voltage-clamp analyses have shown that apamin at low concentrations specifically blocks the Ca2"-dependent K+ channel in differentiated neuroblastoma cells. Binding experiments with highly radiolabeled toxin indicate that the dissociation constant of the apamin-receptor complex in differentiated neuroblastoma cells is 15-22 pM and the maximal binding capacity is 12 fmol/ mg of protein. The receptor is destroyed by proteases, suggesting that it is a protein.The binding capacity ofneuroblastoma cells for radiolabeled apamin dramatically increases during the transition from the nondifferentiated to the differentiated state.

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I3H]Phencyclidine binds to synaptic membranes from rat brain in a saturable, reversible, and selective fashion, with a dissociation constant Kd of 0.25 jiM and a maximal binding capacity of 2.4 pmol/mg of membrane protein-i.e., 250 pmol/g of brain. The binding activity is concentrated in synaptosomal fractions, is higher in cerebral cortex and corpus striatum than in other parts of the rat brain, and is not detectable in the spinal cord. Only molecules of the phencyclidine series and ketamine are able to bind to the phencyclidine receptor.[3H]Phencyclidine bound to its receptor is not displaced by the classical neurotransmitters or neuromodulators. There is a good correlation between the apparent affinities of a series of phencyclidine analogs for the phencyclidine receptor and the pharmacological activities of these analogs as measured by the rotarod assay.Phencyclidine [N-(l-phenylcyclohexyl)piperidine] was introduced in the late 1950s as an intravenous general anesthetic that was nontoxic and nonflammable and produced minimal cardiorespiratory depression (1, 2). These clinical advantages were unfortunately offset by its prolonged duration of action and psychotomimetic effects (3), properties that have contributed to the emergence of phencyclidine as a major drug of abuse in the United States (4, 5). Phencyclidine produces long-lasting psychosis thought to resemble schizophrenia more than does the psychosis produced by any other psychotomimetic (3). Phencyclidine exaggerates psychopathology. Schizophrenics experience severe thought disorders and behavioral problems as long as 1 month after a single injection. Quiet mental patients become catatonic, and reactive ones become overreactive and restless (6, 7). In this paper we investigate the characterization of the phencyclidine receptor in mammalian brain. The data presented here show that a component present in rat brain membranes binds [3H]phencyclidine specifically and with a fairly high affinity. This component may be the physiologically important receptor that mediates the effects of phencyclidine in the central nervous system. MATERIALS AND METHODSMaterials. Molecules in the phencyclidine series were synthesized as previously described (8). The structures of these drugs are shown in Table 1. [3H]Phencyclidine was prepared on request at New England Nuclear by catalytic reduction at room temperature with tritium gas and purification of the reaction mixture on a silica gel column by high-pressure liquid chromatography. The purity of the product was established by thin-layer chromatography on silica gel in either methanol or 1-butanol/acetic acid/water, 25:4:10 vol/vol/vol. The specific radioactivity of the pure [3H]phencyclidine was 60 Ci/mmol (1 Ci = 3.7 x 1010 becquerels). Trans -r2OMt * Ph, phenyl; Th, 2-thienyl. 4678The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

An in vivo fluorescent deltorphin (Fluo-DLT) internalization assay was used to assess the distribution and regulation of pharmacologically available ␦ opioid receptors (␦ORs) in the rat lumbar (L4 -5) spinal cord. Under basal conditions, intrathecal injection of Fluo-DLT resulted in the labeling of numerous ␦OR-internalizing neurons throughout dorsal and ventral horns. The distribution and number of Fluo-DLT-labeled perikaryal profiles were consistent with that of ␦OR-expressing neurons, as revealed by in situ hybridization and immunohistochemistry, suggesting that a large proportion of these cells was responsive to intrathecally administered ␦OR agonists. Pretreatment of rats with morphine for 48 hr resulted in a selective increase in Fluo-DLT-labeled perikaryal profiles within the dorsal horn. These changes were not accompanied by corresponding augmentations in either ␦OR mRNA or 125 I-deltorphin-II binding levels, suggesting that they were attributable to higher densities of cell surface ␦OR available for internalization rather than to enhanced production of the receptor. Unilateral dorsal rhizotomy also resulted in increased Fluo-DLT internalization in the ipsilateral dorsal horn when compared with the side contralateral to the deafferentation or to non-deafferented controls, suggesting that ␦OR trafficking in dorsal horn neurons may be regulated by afferent inputs. Furthermore, morphine treatment no longer increased Fluo-DLT internalization on either side of the spinal cord after unilateral dorsal rhizotomy, indicating that OR-induced changes in the cell surface availability of ␦OR depend on the integrity of primary afferent inputs. Together, these results suggest that regulation of ␦OR responsiveness through OR activation in this region is linked to somatosensory information processing.