C J Shelton’s research while affiliated with Cardiff University and other places

The effect of hyperbaric pressure on the inhibitory glycine receptor has been investigated in voltage-clamped Xenopus oocytes microinjected with cRNA encoding the human alpha-1 glycine receptor subunit. Heterologous expression of the human alpha-1 subunit generated functional glycine-gated channels with properties typical of native receptors. Glycine elicited a concentration-dependent inward current which reversed polarity at -25 mV and was antagonised by nanomolar concentrations of strychnine. Concentration-response curves established for the homomeric alpha-1 glycine receptor at 5, 10 and 15 MPa were progressively shifted to the right with respect to the concentration response curve established at atmospheric pressure (0.1 MPa). Pressure had no effect on the maximal response. The EC(50) values at 0.1, 5, 10 and 15 MPa were 190 mu M, 222 mu M, 338 mu M and 482 mu M, respectively. The results demonstrate that a receptor comprised solely of the human alpha-subunit is sensitive to pressure in the range that affects divers and at which the native rat spinal cord receptor is affected. This finding is discussed in the context of the postulated binding sites for glycine and the implications for the design of drugs to protect divers from the effects of pressure.

The effect of high pressure on the response to glycine or kainate of voltage-clamped Xenopus oocytes micro-injected with messenger-RNA derived from either rat spinal cord or whole brain, respectively, has been investigated. Current responses were measured at 1 bar (= 10(5) Pa), 50 bar, 100 bar and 150 bar, with PO2 fixed at 1 bar and the balance helium. Glycine elicited a depolarizing current response which was antagonized by nanomolar concentrations of strychnine. The responses reversibly desensitized, with a decay constant of 0.01 s-1, when glycine concentrations greater than 250 microM were used. The decay constant was insensitive to both glycine concentration and pressure. Resensitization was complete within 4 min. Kainate elicited a depolarizing current which was non-desensitizing. The response was slightly sensitive to glutamate diethyl ester (50 microM), which increased the EC50 by 25%. The action of glycine was highly pressure sensitive. The dose-response curves established at 50 bar, 100 bar and 150 bar were shifted progressively to the right, with no effect on the maximal current. The EC50 increased from 216 microM to 296 microM at 50 bar, to 345 microM at 100 bar, and to 425 microM at 150 bar. The action of kainate was unaffected by pressure. No shift in the dose-response curves was established, nor was there any effect on the maximum current. The EC50 was 113 microM at 1 bar, and 111 microM at both 50 bar and 100 bar.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of general anesthetics and pressure on receptors from the mammalian central nervous system have been investigated using oocyte expression techniques. Poly A+ mRNA extracted from rat whole brain was injected into mature Xenopus oocytes producing depolarizing responses to the fast excitatory neurotransmitters NMDA and kainate and the inhibitory neurotransmitters GABA and glycine. An apparatus was constructed to allow agonist dose-response curves to be determined at high pressures using voltage-clamped oocytes. This was used to investigate the excitatory transmitter kainate. It was found that anesthetics depress the current induced by kainate whereas pressure does not appear to affect the responses associated with this transmitter. Furthermore it was found that pressure does not reverse (or modify in any way) the changes in response brought about by application of anesthetics.