No abstract

We hypothesized that TRPV4, a member of the transient receptor family of ion channels, functions as a sensory transducer for osmotic stimulus-induced nociception. We found that, as expected for a transducer molecule, TRPV4 protein is transported in sensory nerve distally toward the peripheral nerve endings. In vivo single-fiber recordings in rat showed that hypotonic solution activated 54% of C-fibers, an effect enhanced by the hyperalgesic inflammatory mediator prostaglandin E2. This osmotransduction causes nociception, since administration of a small osmotic stimulus into skin sensitized by PGE2 produced pain-related behavior. Antisense-induced decrease in expression of TRPV4 confirmed that the channel is required for hypotonic stimulus-induced nociception. Thus, we conclude that TRPV4 can function as an osmo-transducer in primary afferent nociceptive nerve fibers. Because this action is enhanced by an inflammatory mediator, TRPV4 may be important in pathological states and may be an attractive pharmacological target for the development of novel analgesics.

Voriconazole is a new antifungal agent effective in the treatment of invasive aspergillosis. Interpatient variation in plasma concentrations is considerable--more than 100-fold. We describe 3 patients with diverse manifestations of toxicity (e.g., hallucinations, hypoglycemia, electrolyte disturbance, and pneumonitis) possibly attributable to high voriconazole concentrations. Measurement of plasma concentrations could be helpful in optimizing voriconazole dosages.

It has been shown that the analgesic and cyclooxygenase inhibitor activity of ketorolac tromethamine (KT), which is marketed as the racemic mixture of (-)S and (+)R enantiomers, resides primarily with (-)S ketorolac and that the ulcerogenic activity of this agent also resides in (-)S ketorolac. Resolution of individual enantiomers for analysis in plasma samples has been accomplished by two methods: derivatization to form diastereomers that are separated by HPLC, or direct HPLC using a chiral phase column. When mice and rats were given oral solutions of (-)S and (+) KT, it was found that the kinetics and interconversion of the enantiomers were species and dose dependent. Interconversion was higher in mice than in rats; when (-)S KT was administered, 71% of the area under the concentration-time curve (AUC) was due to (+)R ketorolac in mice, compared with 12% in rats. More interconversion was observed at higher doses; the percent of AUC due to (-)S ketorolac when (+)R KT was administered increased from 12% to 25% in mice and from 2% to 8% in rats. In general, more interconversion occurred from (-)S to (+)R ketorolac in the animal studies. Human subjects were given single oral solution doses of racemic KT (30 mg), (-)S KT (15 mg), and (+)R KT (15 mg). The plasma concentrations of (-)S ketorolac were lower than (+)R ketorolac at all sample times after racemic KT (22% of the AUC was due to (-)S ketorolac). When (+)R KT was administered, (-)S ketorolac was not detectable and interconversion was essentially 0%. When (-)S KT was administered, significant levels of (+)R ketorolac were detectable and interconversion was 6.5%. After all doses, plasma half-life was shorter and clearance greater for (-)S ketorolac than for (+)R ketorolac. Thus, in humans very little or no interconversion of (+)R to (-)S was observed, and interconversion of (-)S to (+)R was minimal (6.5%). These data demonstrate that the kinetics and interconversion of the enantiomers of ketorolac is different in animals and humans as well as from most other NSAIDs. This may be due to more rapid excretion or metabolism of (-)S ketorolac and a different mechanism of interconversion.

To ascertain the anionic sites on the nicotinic receptor to which acetylcholine and other quaternary ammonium ligands bind, we have examined the role of an aspartyl residue (Asp-152) in the ␣-subunit. Prior photolytic labeling with agonist analogues of the neighboring residues Trp-149 and Tyr-151 suggests that their side chains reside on the binding face (also termed the (؉)-or counterclockwise face) of the ␣-subunit. Asp-152 presents an anionic charge in the vicinity of these aromatic residues. Modification of the aspartate to asparagine (D152N) creates a glycosylation signal (Asn-152-GlySer), and we find, on the basis of altered electrophoretic migration, that glycosylation occurs at this position upon cotransfection of the mutant ␣-subunit with ␤-, ␥-, and ␦-subunits. Glycosylation results in a reduction in the capacity of the receptor to assemble; this reduction is manifest in the initial step of dimer formation between the ␣␥-and ␣␦-subunits. The ␣-subunit mutant receptor reaching the assembled pentamer exhibits an altered selectivity for certain ligands. Little reduction in ␣-bungarotoxin binding is observed, whereas affinities for agonists and competitive alkaloid antagonists are reduced substantially. Separation of the contributions of charge removal and glycosylation addition shows that both factors affect agonist affinity, with the charge influence being far more predominant. These findings raise the possibility that a component of the coulombic attraction stabilizing the binding of agonists comes from the aspartyl residue at position 152 in the ␣-subunit.