Mechanosensory channels of sensory cells mediate the sensations of hearing, touch, and some forms of pain. The TRPA1 (a member of the TRP family of ion channel proteins) channel is activated by pain-producing chemicals, and its inhibition impairs hair cell mechanotransduction. As shown here and previously, TRPA1 is expressed by hair cells as well as by most nociceptors (small neurons of dorsal root, trigeminal, and nodose ganglia) and localizes to their sensory terminals (mechanosensory stereocilia and peripheral free nerves, respectively). Thus, TRPA1 channels are proposed to mediate transduction in both hair cells and nociceptors. Accordingly, we find that heterologously expressed TRPA1 display channel behaviors expected for both auditory and nociceptive transducers. First, TRPA1 and the hair cell transducer share a unique set of pore properties not described for any other channel (block by gadolinium, amiloride, gentamicin, and ruthenium red, a ranging conductance of ϳ100 pS that is reduced to 54% by calcium, permeating calcium-induced potentiation followed by closure, and reopening by depolarization), supporting a direct role of TRPA1 as a pore-forming subunit of the hair cell transducer. Second, TRPA1 channels inactivate in hyperpolarized cells but remain open in depolarized cells. This property provides a mechanism for the lack of desensitization, coincidence detection, and allodynia that characterize pain by allowing a sensory neuron to respond constantly to sustained stimulation that is suprathreshold (i.e., noxious) and yet permitting the same cell to ignore sustained stimulation that is subthreshold (i.e., innocuous). Our results support a TRPA1 role in both nociceptor and hair cell transduction.

Acad. Sci. USA (97, 13913-13918; First Published November 28, 2000; 10.1073͞pnas.250478897), the authors note that the exponents of some entries in Table 1 were misprinted. The correct values appear below. www.pnas.org͞cgi͞doi͞10.1073͞pnas.191384698 STATISTICS, GENETICS. For the article ''Significance analysis of microarrays applied to the ionizing radiation response'' by Virginia Goss Tusher, Robert Tibshirani, and Gilbert Chu, which appeared in number 9, April 24, 2001, of Proc. Natl. Acad. Sci. USA (98, 5116-5121; First Published April 17, 2001; 10.1073͞pnas.091062498), the authors note the following: ''In our discussion of the pairwise fold change method on page 5118, we cited a paper by Ly et al., crediting them for the method. We did not mean to imply that it was deficient for the analysis of their experiments. In fact, Ly et al. incorporate (98,(6384)(6385)(6386)(6387)(6388)(6389), the authors wish to correct the position given for the amino acid that was mutated in the patient. The mutation ''R187W'' should be ''R188W. '' www.pnas.org͞cgi͞doi͞10.1073͞pnas.191390798 FEB2, 19p; FEB3, and FEB4,. A small population of individuals with FS has additional generalized epilepsy (1) or afebrile seizures. Genes for a ␤-subunit (␤1) and an ␣ I -subunit (Na v 1.1: SCN1A) (10) of the neuronal voltage-gated Na ϩ channel have been identified to be responsible for generalized epilepsy with febrile seizures plus (GEFSϩ) type 1 and 2, respectively (11, 12). However, a large number of patients with GEFSϩ still show no mutation for those genes. These, therefore, suggest that other genes might also be involved in GEFSϩ and FS associated with afebrile seizures. The chromosomal locus 2q24, in which GEFSϩ has been mapped, harbors not only Na v 1.1 but also other ␣-subunits including Na v 1.2 (SCN2A) (10,(13)(14)(15). Given that Na v 1.2 is also expressed in high levels in the central nervous system with a tissue-specific profile (16), Na v 1.2 is an intriguing candidate. In the present study, we report a mutation of Na v 1.2 found in a patient with FS and afebrile seizures. A channel harboring the mutation shows abnormal electrophysiological properties that may underlie the neuronal hyperexcitability that triggers seizure activity. Materials and MethodsPatients and Pedigrees. This study recruited nineteen unrelated Japanese families with members clinically diagnosed with GEFSϩ or febrile seizures associated with afebrile seizures. Each participating subject or a responsible adult signed an informed consent form approved by the Ethics Review Committee of Fukuoka University or similar committees of the participating institutions. The proband of family K1 is a 6-yr-old boy with normal development (Fig. 1A). He had the first febrile seizure (FS) at 8 months of age and suffered 17 episodes of FS thereafter at both high and low grade fever. The FS were generalized tonic or tonic-clonic convulsions with duration of 1-5 min per episode. Since 4 yr of age, he also has experienced brief afebrile atonic seizures 5 times. The...

Varitint-waddler (Va andVaA very common cause of deafness is the loss of hair cells, which degenerate because of environmental factors or genetic mutations. Varitint-waddler (Va) mice have a mutation that causes an alanine-to-proline substitution (A419P) in the fifth transmembrane domain of TRPML3, a presumed ion channel. A second allele, Va J , contains the A419P mutation in cis to an I362T mutation (1). The earliest sign of inner ear damage in Va mice is hair cell degeneration, which begins in embryogenesis as hair cells differentiate and continues postnatally. During degeneration, the hair cells bulge out of the apical side of the epithelium and become extruded. Eventually these mice also develop abnormalities in supporting cells, in the tectorial membrane, and in the stria vascularis and in the endocochlear potential that they help generate. The defects of Va J mice are similar although less severe, suggesting that the mutation I362T attenuates the effects of A419P (1-3). A comprehensive examination of TRPML3 expression in inner ear has not been published. Expression of TRPML3 protein in hair cells was suggested by immunocytochemical methods, although results for other inner ear cell types were not reported (1).Mutations in two other TRP channels, Drosophila TRP and human TRPML1 (mutations in which cause mucolipidosis type IV), are associated with degeneration of the retina. In both cases, the channels seem necessary for cellular viability, because recessive, loss-of-function mutations cause degeneration (4, 5).By contrast, the Va and Va J mutations in TRPML3 are semidominant, although it remains unclear whether this is due to gain-of-function, haploinsufficiency, or dominant-negative effects (1, 6). ResultsWe first established which cells expressed TRPML3 in the inner ear of wild-type mice. In situ hybridization on mouse inner ear using probes from two nonoverlapping regions of TRPML3 mRNA gave identical results [ Fig. 1 and supporting information (SI) Fig. 6]. TRPML3 mRNA was most abundant in the marginal cells of the cochlear stria vascularis and the dark cells of the vestibule (Fig. 1 i, l, and m and SI Fig. 6 i, l, and m). Both cell types are involved in producing the endolymph and therefore the endocochlear potential (7-9). This expression pattern is in keeping with the reduced stria vascularis and the rounding up and loss of the cytoplasmic processes of its marginal cells in Va/Va and Va/ϩ mice (2) and with the reduced endocochlear potentials of the Va J mutants (1, 3).In addition, other cells that line the cochlear scala media and the connected vestibular endolymphatic spaces expressed TRPML3 mRNA, including (i) supporting cells at the marginal part of the lesser epithelial ridge (Hensen and Claudius cells), which in Va and Va J mice (homozygotes and heterozygotes) appear undifferentiated or degenerate ( Fig. 1i and SI Fig. 6i); (ii) cells of the spiral limbus that produce the tectorial membrane, which in Va mice develops abnormally and fails to attach to the reticular lamina ( Fig. 1i and SI Fig. 6i...

Neurons with the capacity to discharge at high rates--'fast-spiking' (FS) neurons--are critical participants in central motor and sensory circuits. It is widely accepted that K+ channels with Kv3.1 or Kv3.2 subunits underlie fast, delayed-rectifier (DR) currents that endow neurons with this FS ability. Expression of these subunits in heterologous systems, however, yields channels that open at more depolarized potentials than do native Kv3 family channels, suggesting that they differ. One possibility is that native channels incorporate a subunit that modifies gating. Molecular, electrophysiological and pharmacological studies reported here suggest that a splice variant of the Kv3.4 subunit coassembles with Kv3.1 subunits in rat brain FS neurons. Coassembly enhances the spike repolarizing efficiency of the channels, thereby reducing spike duration and enabling higher repetitive spike rates. These results suggest that manipulation of K3.4 subunit expression could be a useful means of controlling the dynamic range of FS neurons.

No abstract

1 Anisatin, a toxic, insecticidally active component of Sikimi plant, is known to act on the GABA system. In order to elucidate the mechanism of anisatin interaction with the GABA system, wholecell and single-channel patch clamp experiments were performed with rat dorsal root ganglion neurons in primary culture. 2 Repeated co-applications of GABA and anisatin suppressed GABA-induced whole-cell currents with an EC 50 of 1.10 mM. No recovery of currents was observed after washout with anisatin-free solution.3 However, pre-application of anisatin through the bath had no eect on GABA-induced currents. The decay phase of currents was accelerated by anisatin. These results indicate that anisatin suppression of GABA-induced currents requires opening of the channels and is use-dependent. 4 Anisatin suppression of GABA-induced currents was not voltage dependent. 5 Picrotoxinin attenuated anisatin suppression of GABA-induced currents. [ 3 H]-EBOB binding to rat brain membranes was competitively inhibited by anisatin. These data indicated that anisatin bound to the picrotoxinin site. 6 At the single-channel level, anisatin did not alter the open time but prolonged the closed time. The burst duration was reduced and channel openings per burst were decreased indicating that anisatin decreased the probability of openings.

No abstract

Whereas the selective toxicity of insecticides between insects and mammals has a long history of studies, it is now becoming abundantly clear that, in many cases, the differential action of insecticides on insects and mammalian target receptor sites is an important factor. In this paper, we first introduce the mechanism of action and the selective toxicity of pyrethroids as a prototype of study. Then, a more detailed account is given for fipronil, based primarily on our recent studies. Pyrethroids keep the sodium channels open for a prolonged period of time, causing elevation of the depolarizing after-potential. Once the after-potential reaches the threshold for excitation, repetitive after-discharges are produced, resulting in hyperexcitation of intoxicated animals. Only about 1% of sodium channels needs to be modified to produce hyperexcitation, indicating a high degree of toxicity amplification from sodium channels to animals. Pyrethroids were >1000-fold more potent on cockroach sodium channels than rat sodium channels, and this forms the most significant factor to explain the selective toxicity of pyrethroids in insects over mammals. Fipronil, a phenylpyrazole, is known to act on the γ-aminobutyric acid receptor to block the chloride channel. It is effective against certain species of insects that have become resistant to most insecticides, including those acting on the γ-aminobutyric acid receptor, and is much more toxic to insects than to mammals. Recently, fipronil has been found to block glutamate-activated chloride channels in cockroach neurons in a potent manner. Since mammals are devoid of this type of chloride channel, fipronil block of the glutamate-activated chloride channel is deemed responsible, at least partially, for the higher selective toxicity to insects over mammals and for the lack of cross-resistance. Human & Experimental Toxicology (2007) 26, 361-366