Donatella Tombaccini

Effects of glucose on insulin release and 86Rb permeability in cultured neonatal and adult rat islets

Glucose‐induced insulin release and modifications in 86Rb outflow were studied in cultured neonatal and adult rat islets. The dose‐response curve for neonatal islets was steeper than for adult islets and the maximal response was clearly shifted towards lower glucose concentrations. In neonatal islets, glucose‐induced insulin release was inhibited by the Ca2+‐channel blocker, nifedipine. In the absence of glucose, the 86Rb outflow from neonatal islets was lower than from adult islets. Also, the glucose‐induced reduction in 86Rb outflow was less pronounced in neonatal islets. Altered K+ permeability in the B‐cell membrane could explain the change in glucose sensitivity of neonatal islets.

A goldfish model for evaluation of the neurotoxicity of ω-conotoxin GVI A and screening of monoclonal antibodies

The neurotoxicity of omega-conotoxin (omega-CgTx), a potent neuronal voltage-sensitive calcium channel blocker, was measured using a new bioassay. omega-CgTx was administered intraperitoneally (ip) to goldfish weighing approximately 1.6 g, and dose-related changes were observed over a 2-hr period. omega-CgTx induced time- and dose-dependent abnormal swimming behavior (ASB) and mortality. The antitoxin activity of the antibodies was investigated in vivo by either (1) preincubation of the antibody with omega-CgTx at 4 degrees C overnight, or (2) pretreatment with antibody, 30 min before omega-CgTx injection in a 10:1 antibody/omega-CgTx molar ratio. The LD50 dose of omega-CgTx in goldfish was 5 nmol/kg ip, and preincubation of monoclonal antibody (50 nmol/kg ip) with omega-CgTx (5 nmol/kg ip) significantly (p less than 0.05) reduced mortality, ASB, and toxicity time. The antitoxin activity of the monoclonal antibodies evidenced in the goldfish bioassay was further tested in the conscious rat. In the rat, the increases in mean arterial pressure and heart rate induced by omega-CgTx (0.03 nmol/rat icv) were significantly (p less than 0.02 and p less than 0.01, respectively) attenuated by preincubation of the toxin with the antibody (0.3 nmol/rat). We conclude that the goldfish bioassay provides a simple, accurate, and inexpensive in vivo model for the study of the toxicity of omega-CgTx.

Monoclonal antibodies against the presynaptic calcium channel antagonist ω-conotoxin GVI A from cone snail poison

Monoclonal antibodies have been prepared against co‐conotoxin GVI A, a peptide isolated from marine snails of the genus Conus (Conus geographus and Conus magus). This toxin is a blocker of select presynaptic Ca2+ channels in the central nervous system. Antigenic ω‐conotoxin GVI A was synthesized as a covalent conjugate with bovine serum albumin and injected s.c. An ELISA assay combined with a competitive inhibition assay was used to select and characterize monoclonal antibodies able to recognize and bind the free toxin. Several of the antibodies were found to block ω‐conotoxin GVI A inhibition of 45Ca transport into rat brain synaptosomes and to block ω‐conotoxin GVI A binding to membranes from the same preparation. The antibodies recognize native, synthetic toxin, and are useful for analysis of toxin in biological fluids.

Differences in K+ permeability between cultured adult and neonatal rat islets of Langerhans in response to glucose, tolbutamide, diazoxide, and theophylline.

The effects of glucose, tolbutamide, and diazoxide on K+ permeability in neonatal and adult rat pancreatic islets, maintained in culture 1 week, were investigated by measuring the 86Rb outflow rate from prelabeled islets. In the absence of glucose, the 86Rb efflux was significantly lower in neonatal than adult islets. Raising the glucose concentration to 2.8, 5.6, 8.3, and 11.1 mM produced a marked reduction in the 86Rb efflux in adult islets but only a minor reduction in neonatal islets. The effect of tolbutamide to reduce, and diazoxide to increase, the 86Rb efflux was also less in neonatal islets. These results are discussed with respect to previously reported differences in insulin secretion from neonatal and adult islets in culture.

Monoclonal Antibodies and the Thyrotropin Receptor

Thyrotropin (TSH) is a pituitary glycoprotein hormone whose primary role is to regulate thyroid cell function (Dumont, 1971; Field, 1980; Robbins et al., 1980; Kohn et al., 1983). The interaction of TSH with a specific receptor on the thyroid cell surface induces changes in adenylate cylcase activity which result in the following tissue responses: enhanced iodide uptake, thyroglobulin biosynthesis, iodination of thyroglobulin, degradation of iodinated thyroglobulin to form thyroid hormone, and the release of thyroid hormone (T3 and T4) into the bloodstream. Although changes in all of the above activities have been and can be used to define receptor function, definition of the structure of the TSH receptor on a molecular level, as with all other receptors, has required the identification of specific membrane components using binding studies and 125I-labeled hormone (Kohn, 1978; Kohn and Shifrin, 1982; Kohn et al., 1983).