Vanilloids such as capsaicin have algesic properties and seem to mediate their effects via activation of the vanilloid receptor 1 (VR1), a ligand-gated ion channel highly expressed on primary nociceptors. Although blockade of capsaicin-induced VR1 activation has been demonstrated in vitro and in vivo with the antagonist capsazepine, efficacy in rat models of chronic pain has not been observed with this compound. Here, we describe the in vitro pharmacology of a highly potent VR1 antagonist, N-(4-tertiarybutylphenyl)-4-(3-chloropyridin-2-yl)tetrahydropyrazine-1(2H)-carbox-amide (BCTC). Similar to capsazepine, this compound inhibits capsaicin-induced activation of rat VR1 with an IC 50 value of 35 nM. Interestingly however, BCTC also potently inhibits acid-induced activation of rat VR1 (IC 50 value of 6.0 nM), whereas capsazepine is inactive. Similarly, in the rat skin-nerve preparation both BCTC and capsazepine block capsaicin-induced activation, whereas the response to acidification is inhibited by BCTC, but not by capsazepine. Specificity for VR1 was demonstrated against 63 other receptor, enzyme, transporter, and ion channel targets. BCTC was orally bioavailable in the rat, demonstrating a plasma half-life of ϳ1 h and significant penetration into the central nervous system. Thus, BCTC is a high potency, selective VR1 antagonist that, unlike capsazepine, has potent blocking effects on low pH-induced activation of rat VR1. These properties make it a more suitable candidate than capsazepine for testing the role played by VR1 in rat models of human disease.

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Abstract. We have transformed Drosophila melanogaster with a genomic construct containing the entire wild-type myosin heavy-chain gene, Mhc, together with •9 kb of flanking DNA on each side. Three independent lines stably express myosin heavy-chain protein (MHC) at approximately wild-type levels. The MHC produced is functional since it rescues the mutant phenotypes of a number of different Mhc alleles: the amorphic allele Mhc 1, the indirect flight muscle and jump muscle-specific amorphic allele Mhc ~°, and the hypomorphic allele Mhc 2. We show that the Mhc ~ mutation is due to the insertion of a transposable element in an intron of Mhc.Since a reduction in MHC in the indirect flight muscles alters the myosin/actin protein ratio and results in myofibrillar defects, we determined the effects of an increase in the effective copy number of Mhc. The presence of four copies of Mhc results in overabundance of the protein and a flightless phenotype. Electron microscopy reveals concomitant defects in the indirect flight muscles, with excess thick illaments at the periphery of the myofibrils. Further increases in copy number are lethal. These results demonstrate the usefulness and potential of the transgenic system to study myosin function in Drosophila. They also show that overexpression of wild-type protein in muscle may disrupt the function of not only the indirect flight but also other muscles of the organism.

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4‐(4‐Fluorophenoxy)benzaldehyde semicarbazone (V102862) was initially described as an orally active anticonvulsant with robust activity in a variety of rodent models of epilepsy. The mechanism of action was not known. We used whole‐cell patch‐clamp techniques to study the effects of V102862 on native and recombinant mammalian voltage‐gated Na+ channels. V102862 blocked Na+ currents (INa) in acutely dissociated cultured rat hippocampal neurons. Potency increased with membrane depolarization, suggesting a state‐dependent mechanism of inhibition. There was no significant effect on the voltage dependence of activation of INa. The dissociation constant for the inactivated state (KI) was ∼0.6 μM, whereas the dissociation constant for the resting state (KR) was >15 μM. The binding to inactivated channels was slow, requiring a few seconds to reach steady state at −80 mV. The mechanism of inhibition was characterized in more detail using human embryonic kidney‐293 cells stably expressing rat brain type IIA Na+ (rNav1.2) channels, a major Na+ channel α subunit in rat hippocampal neurons. Similar to hippocampal neurons, V102862 was a potent state‐dependent blocker of rNav1.2 channels with a KI of ∼0.4 μM and KR ∼30 μM. V102862 binding to inactivated channels was relatively slow (k+≃1.7 μM−1 s−1). V102862 shifted the steady‐state availability curve in the hyperpolarizing direction and significantly retarded recovery of Na+ channels from inactivation. These results suggest that inhibition of voltage‐gated Na+ channels is a major mechanism underlying the anticonvulsant properties of V102862. Moreover, understanding the biophysics of the interaction may prove to be useful in designing a new generation of potent Na+ channel blocker therapeutics. British Journal of Pharmacology (2005) 144, 801–812. doi:10.1038/sj.bjp.0706058