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Takashi Kurahashi

Publications and source records attributed to Takashi Kurahashi.

10 recordsLinked to original sources
PubMed2006

Odorant inhibition of the olfactory cyclic nucleotide-gated channel with a native molecular assembly.

Human olfaction comprises the opposing actions of excitation and inhibition triggered by odorant molecules. In olfactory receptor neurons, odorant molecules not only trigger a G-protein-coupled signaling cascade but also generate various mechanisms to fine tune the odorant-induced current, including a low-selective odorant inhibition of the olfactory signal. This wide-range olfactory inhibition has been suggested to be at the level of ion channels, but definitive evidence is not available. Here, we report that the cyclic nucleotide-gated (CNG) cation channel, which is a key element that converts odorant stimuli into electrical signals, is inhibited by structurally unrelated odorants, consistent with the expression of wide-range olfactory inhibition. Interestingly, the inhibitory effect was small in the homo-oligomeric CNG channel composed only of the principal channel subunit, CNGA2, but became larger in channels consisting of multiple types of subunits. However, even in the channel containing all native subunits, the potency of the suppression on the cloned CNG channel appeared to be smaller than that previously shown in native olfactory neurons. Nonetheless, our results further showed that odorant suppressions are small in native neurons if the subsequent molecular steps mediated by Ca(2+) are removed. Thus, the present work also suggests that CNG channels switch on and off the olfactory signaling pathway, and that the on and off signals may both be amplified by the subsequent olfactory signaling steps.

Animals↗
PubMed2005

Mechanism of signal amplification in the olfactory sensory cilia.

Molecular mechanisms underlying olfactory signal amplification were investigated by monitoring cAMP dynamics in the intact sensory cilia. We saw that [cAMP]i increased superlinearly with time during odorant stimuli for >1 s. This time course was remarkably different from that obtained with the rapid quench method previously applied to the in vitro preparation, in which [cAMP]i change has been reported to be transient. The superlinear increase of [cAMP]i was attributable to a gradual increase of cAMP production rate that was consistent with the thermodynamical interaction model between elemental molecules, as has been revealed on the rod photoreceptor cell. It thus seems likely that the fundamental mechanism for molecular interactions between olfactory transduction elements is similar to that of the rod. In olfaction, however, cAMP production was extremely small (approximately 200,000 molecules/s/cell at the maximum), in contrast to the cGMP hydrolysis in the rod (250,000 molecules/photon). The observed numbers indicate that the olfactory receptor cell has lower amplification at the enzymatic cascade. Seemingly, such low amplification is a disadvantage for the signal transduction, but this unique mechanism would be essential to reduce the loss of ATP that is broadly used for the activities of cells. Apparently, transduction by a smaller number of second-messenger formations would be achieved by the fine ciliary structure that has a high surface-volume ratio. In addition, it is speculated that this low amplification at their enzymatic processes may be the reason why the olfactory receptor cell has acquired high amplification at the final stage of transduction channels, using Ca2+ as a third messenger.

Adenylyl Cyclases↗
PubMed2004

Mechanisms determining the dynamic range of the bullfrog olfactory receptor cell.

Spike discharges of single olfactory receptor cells (ORCs) were recorded with the whole cell patch-clamp method applied to slice preparation. In parallel, activities of transduction channels were measured under the voltage-clamp condition. When cells were stimulated by odorants, 54 out of 306 cells exhibited inward current responses (10 mM cineole in the puffer pipette). The amplitude of the inward current was dependent on the stimulus period, reflecting the time integration for the stimulus dose, and the relation could be fitted by the Hill equation. Under the current-clamp condition, current injection induced spike discharges. In cells showing repetitive firings, the firing frequency was dependent on the amount of injected current. The relation was fitted by the Michaelis-Menten equation showing saturation. When cells were responsive to the odorant and had abilities to discharge repetitive spikes, the depolarizing responses were accompanied by repetitive spikes. In those cells, the spike frequency was dose-dependent, expressing saturation similar to the result obtained by current injection. Since both transduction channel and spike generative steps expressed saturation in their dose dependences, we explored what step(s) actually determines saturation in ORC signaling processes. By examining dose-response relations of both the current and spikes in the same cells, saturating dose was found to be dependent largely on that of the transduction step. This suggests that the dynamic range is fundamentally determined by the transduction system. In addition, a simple model derived from the nonlinearity of the plasma membrane could explain that a critical level of dynamic range was, at least in part, modified by the membrane nonlinearity.

Action Potentials↗
PubMed2004

Bhc-cNMPs as either water-soluble or membrane-permeant photoreleasable cyclic nucleotides for both one- and two-photon excitation.

Cyclic nucleoside monophosphates (cNMPs) play key roles in many cellular regulatory processes, such as growth, differentiation, motility, and gene expression. Caged derivatives that can be activated by irradiation could be powerful tools for studying such diverse functions of intracellular second messengers, since the spatiotemporal dynamics of these molecules can be controlled by irradiation with appropriately focused light. Here we report the synthesis, photochemistry, and biological testing of 6-bromo-7-hydroxycoumarin-4-ylmethyl esters of cNMP (Bhc-cNMP) and their acetyl derivatives (Bhc-cNMP/Ac) as new caged second messengers. Irradiation of Bhc-cNMPs quantitatively produced the parent cNMPs with one-photon uncaging efficiencies (Phiepsilon) of up to one order of magnitude better than those of 2-nitrophenethyl (NPE) cNMPs. In addition, two-photon induced photochemical release of cNMP from Bhc-cNMPs (7 and 8) can be observed with the two-photon uncaging action cross-sections (delta(u)) of up to 2.28 GM (1 GM=10(-50) cm(4) s photon(-1)), which is the largest value among those of the reported Bhc-caged compounds. The wavelength dependence of the delta(u) values of 7 revealed that the peak wavelength was twice that of the one-photon absorption maximum. Bhc-cNMPs showed practically useful water solubility (nearly 500 microM), whereas 7-acetylated derivatives (Bhc-cNMPs/Ac) were expected to have a certain membrane permeability. Their advantages were demonstrated in two types of biological systems: the opening of cAMP-mediated transduction channels in newt olfactory receptor cells and cAMP-mediated motility responses in epidermal melanophores in scales from medaka fish. Both examples showed that Bhc and Bhc/Ac caged compounds have great potential for use in many cell biological applications.

Animals↗
PubMed2003

Cross-adaptation between olfactory responses induced by two subgroups of odorant molecules.

It has long been believed that vertebrate olfactory signal transduction is mediated by independent multiple pathways (using cAMP and InsP3 as second messengers). However, the dual presence of parallel pathways in the olfactory receptor cell is still controversial, mainly because of the lack of information regarding the single-cell response induced by odorants that have been shown to produce InsP3 exclusively (but not cAMP) in the olfactory cilia. In this study, we recorded activities of transduction channels of single olfactory receptor cells to InsP3-producing odorants. When the membrane potential was held at -54 mV, application of InsP3-producing odorants to the ciliary region caused an inward current. The reversal potential was 0 +/- 7 mV (mean +/- SD, n = 10). Actually, InsP3-producing odorants generated responses in a smaller fraction of cells (lilial, 3.4%; lyral, 1.7%) than the cAMP-producing odorant (cineole, 26%). But, fundamental properties of responses were surprisingly homologous; namely, spatial distribution of the sensitivity, waveforms, I-V relation, and reversal potential, dose dependence, time integration of stimulus period, adaptation, and recovery. By applying both types of odorants alternatively to the same cell, furthermore, we observed cells to exhibit symmetrical cross-adaptation. It seems likely that even with odorants with different modalities adaptation occurs completely depending on the amount of current flow. The data will also provide evidence showing that olfactory response generation and adaptation are regulated by a uniform mechanism for a wide variety of odorants.

Adaptation, Physiological↗
PubMed2003

Identification of second messenger mediating signal transduction in the olfactory receptor cell.

One of the biggest controversial issues in the research of olfaction has been the mechanism underlying response generation to odorants that have been shown to fail to produce cAMP when tested by biochemical assays with olfactory ciliary preparations. Such observations are actually the original source proposing a possibility for the presence of multiple and parallel transduction pathways. In this study the activity of transduction channels in the olfactory cilia was recorded in cells that retained their abilities of responding to odorants that have been reported to produce InsP3 (instead of producing cAMP, and therefore tentatively termed "InsP3 odorants"). At the same time, the cytoplasmic cNMP concentration ([cNMP]i) was manipulated through the photolysis of caged compounds to examine their real-time interactions with odorant responses. Properties of responses induced by both InsP3 odorants and cytoplasmic cNMP resembled each other in their unique characteristics. Reversal potentials of currents were 2 mV for InsP3 odorant responses and 3 mV for responses induced by cNMP. Current and voltage (I-V) relations showed slight outward rectification. Both responses showed voltage-dependent adaptation when examined with double pulse protocols. When brief pulses of the InsP3 odorant and cytoplasmic cNMP were applied alternatively, responses expressed cross-adaptation with each other. Furthermore, both responses were additive in a manner as predicted quantitatively by the theory that signal transduction is mediated by the increase in cytoplasmic cAMP. With InsP3 odorants, actually, remarkable responses could be detected in a small fraction of cells ( approximately 2%), explaining the observation for a small production of cAMP in ciliary preparations obtained from the entire epithelium. The data will provide evidence showing that olfactory response generation and adaptation are regulated by a uniform mechanism for a wide variety of odorants.

Animals↗
PubMed2002

Photolysis of caged cyclic AMP in the ciliary cytoplasm of the newt olfactory receptor cell.

The effects of cyclic nucleotide monophosphate (cNMP) in the ciliary cytoplasm of the olfactory receptor cell were examined by using photolysis of caged cNMP loaded from the whole-cell patch clamp pipette. Illumination of the cilia induced an inward current at -50 mV. The current amplitude was voltage dependent and the polarity was reversed at +10 mV. The amplitude of the light-induced current was dependent on both light intensity and duration. The intensity-response relation was fitted well by the Hill equation with a coefficient (n(H)) of 4.99 +/- 2.66 (mean +/- S.D., n = 19) and the duration-response relation with a coefficient of 4.03 +/- 1.43 (n = 17). The activation time course of adenylyl cyclase was estimated by comparing the light-induced response with the odorant-induced response. Adenylyl cyclase was activated approximately 260 ms later from the onset of the odorant-stimulation. The light-induced current developed very sharply. This could be explained by the sequential openings of cAMP-gated and Ca2+-activated Cl- channels. At +100 mV, where Ca2+ influx is expected to be very small, the current rising phase became less steep. When the cells were stimulated by long steps of either odour or light, the odorant-induced current showed stronger decay than the light-induced response. This observation suggests that the molecular system regulating desensitization is situated upstream of cAMP production.

Adenylyl Cyclases↗
PubMed2002

Human olfactory contrast changes during the menstrual cycle.

Several lines of studies have reported that olfactory perception is influenced by physical and hormonal conditions. In the present study, we investigated changes of olfactory perception during the menstrual cycle of the human. Cyclopentadecanolide vapor was used and its perception intensity was measured by 6-point category scale methods. We especially focused on a novel concept termed "olfactory contrast" that has been just very recently derived from the patch clamp experiment that uses the solitary olfactory receptor cell. The results obtained from 18 trials (15 subjects) showed that olfactory contrast was significantly enhanced at the ovulatory and/or menstrual phases. It is suggested that olfactory contrast, which we defined as a new parameter, provides a useful tool in many kinds of studies exploring the olfactory perceived ability.

Adult↗
PubMedDate not supplied

Asymmetric alpha-alkylation of phenylacetates using 2-alkylamino-2'-hydroxy-1,1'-binaphthyls as chiral auxiliaries.

alpha-Alkylation of 2-phenylacetate ester of 2-alkylamino-2'-hydroxy-1,1'-binaphthyls with various alkyl iodides proceeded with good stereoselectivities. An intramolecular hydrogen bonding between N-H group and the carbonyl oxygen seemed to play an important role for asymmetric induction. The auxiliary was also applicable to Diels-Alder reaction of an acrylate.

Journal Article↗