Subunit-dependent block by isoflurane of wild-type and mutant alpha(1)S270H GABA(A) receptor currents in Xenopus oocytes.

The volatile anesthetic isoflurane both prolongs and reduces the amplitude of GABA-mediated inhibitory postsynaptic currents (IPSCs) recorded in neurons. To explore the latter effect, we investigated isoflurane-induced inhibition of steady-state desensitized GABA currents in Xenopus oocytes expressing wild-type alpha(1)beta(2), alpha(1)beta(2)gamma(2s), mutant alpha(1)(S270H)beta(2) (serine to histidine at residue 270) or alpha(1)(S270H)beta(2)gamma(2s) receptors. The alpha(1) serine 270 site in TM2 (second transmembrane domain of the subunit) is postulated as a binding site for some volatile agents and is critical for positive modulation of sub-maximal GABA responses by isoflurane. For all receptor combinations, at < or =0.6 mM isoflurane (< or =2 minimum alveolar concentration (MAC)) current inhibitions were not pronounced ( approximately 10%) with block reaching half-maximal levels at supraclinical concentrations ( approximately 2 mM isoflurane, 6 MAC). Comparisons with other GABA(A) receptor blockers indicated that isoflurane blocks in a similar manner to picrotoxin, possibly via the pore of the receptor. The extent of isoflurane-induced inhibition was significantly attenuated by inclusion of the gamma(2s)-subunit but was unaffected by introduction of the S270H mutation in the alpha(1)-subunit. In conclusion, isoflurane binds with low affinity and with subunit-specificity to an inhibitory site on the GABA(A) receptor that is distinct from the site that facilitates positive modulation at the extracellular end of the pore.

Molecular properties important for inhaled anesthetic action on human 5-HT3A receptors.

Although inhaled anesthetics have diverse effects on 5-hydroxytryptamine type 3 (5-HT3A) receptors, the mechanism accounting for this diversity is not understood. Studies have shown that modulation of 5-HT3A receptor currents by n-alcohols depends on molecular volume, suggesting that steric interactions between n-alcohols and their binding sites define their action on this receptor. Electrostatic interactions also play an important role in anesthetic action on other ligand-gated receptors. We aimed to determine the contribution of molecular volume and electrostatics in defining volatile anesthetic actions on 5-HT3A receptors. Human 5-HT3A receptors were expressed in, and recorded from, Xenopus oocytes using the two-electrode voltage-clamp technique. The effects of a range of volatile anesthetics, n-alcohols, and nonhalogenated alkanes on submaximal serotonin-evoked peak currents, and full serotonin concentration-response curves were defined. Volatile anesthetics and n-alcohols, but not alkanes, smaller than 0.120 nm3 enhanced submaximal serotonin-evoked peak currents whereas all larger agents reduced currents. Most compounds tested inhibited maximal serotonin-evoked peak currents to varying degrees. However, only agents smaller than 0.120 nm3 shifted the 5-HT3A receptor's serotonin concentration-response curve to the left, whereas larger anesthetics shifted them to the right. Modulation of human 5-HT3A-mediated currents by volatile anesthetics exhibits a dependence on molecular volume consistent with the n-alcohols, suggesting that both classes of agents may enhance 5-HT3A receptor function via the same mechanism. Furthermore, the enhancing but not inhibiting effects of anesthetic compounds on 5-HT3A receptor currents are modulated by electrostatic interactions.

The effects of isoflurane on desensitized wild-type and alpha 1(S270H) gamma-aminobutyric acid type A receptors.

UNLABELLED: gamma-aminobutyric acid type A receptors (GABA(A)-R) mediate synaptic inhibition and meet many pharmacological criteria required of important general anesthetic targets. During synaptic transmission GABA release is sufficient to saturate, maximally activate, and transiently desensitize postsynaptic GABA(A)-Rs. The resulting inhibitory postsynaptic currents (IPSCs) are prolonged by volatile anesthetics like isoflurane. We investigated the effects of isoflurane on maximally activated and desensitized GABA(A)-R currents expressed in Xenopus oocytes. Wild-type alpha(1)beta(2) and alpha(1)beta(2)gamma(2s) receptors were exposed to 600 microM GABA until currents reached a steady-state desensitized level. At clinical concentrations (0.02-0.3 mM), isoflurane produced a dose-dependent enhancement of steady-state desensitized current in alpha(1)beta(2) receptors, an effect that was less apparent in receptors including a gamma(2s)-subunit. When serine at position 270 is mutated to histidine (alpha(1)(S270H)) in the second transmembrane segment of the alpha(1)-subunit, the currents evoked by sub-saturating concentrations of GABA became less sensitive to isoflurane enhancement. In addition, isoflurane enhancements of desensitized currents were greatly attenuated by this mutation and were undetectable in alpha(1)(S270H)beta(2)gamma(2s) receptors. In conclusion, isoflurane enhancement of GABA(A)-R currents evoked by saturating concentrations of agonist is subunit-dependent. The effects of isoflurane on desensitized receptors may be partly responsible for the prolongation of IPSCs during anesthesia. IMPLICATIONS: Isoflurane enhances desensitized gamma-aminobutyric acid type A receptor (GABA(A)-R) currents, an effect that is subunit-dependent and attenuated by a mutation in an alpha(1)-subunit pore residue of the GABA(A)-R. As GABA release at inhibitory synapses is typically saturating, isoflurane modulation of desensitized receptors may be partly responsible for prolongation of inhibitory postsynaptic currents during anesthesia.