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V E Dionne

Publications and source records attributed to V E Dionne.

At least 19 recordsLinked to original sources
PubMed1992

Spatial pattern of receptor expression in the olfactory epithelium.

A PCR-based strategy for amplifying putative receptors involved in murine olfaction was employed to isolate a member (OR3) of the seven-transmembrane-domain receptor superfamily. During development, the first cells that express OR3 appear adjacent to the wall of the telencephalic vesicle at embryonic day 10. The OR3 receptor is uniquely expressed in a subset of olfactory cells that have a characteristic bilateral symmetry in the adult olfactory epithelium. This receptor and its specific pattern of expression may serve a functional role in odor coding or, alternatively, may play a role in the development of the olfactory system.

Amino Acid Sequence↗
PubMed1992

Chemosensory responses in isolated olfactory receptor neurons from Necturus maculosus.

Olfactory receptor neurons were isolated without enzymes from the mudpuppy, Necturus maculosus, and tested for chemosensitivity. The cells responded to odorants with changes in firing frequency and alterations in excitability that were detected with tight-seal patch electrodes using on-cell and whole-cell recording conditions. Chemosensitive cells exhibited two primary response characteristics: excitation and inhibition. Both types of primary response were observed in different cells stimulated by mixtures of amino acids as well as by the single compound L-alanine, suggesting that there may be more than one transduction pathway for some odorants. Using the normal whole-cell recording method, the chemosensitivity of competent cells washed out rapidly; a resistive whole-cell method was used to record odorant responses under current-clamp conditions. In response to chemical stimulation, excitability appeared to be modulated in several different ways in different cells: odorants induced hyperpolarizing or depolarizing receptor potentials, elicited or inhibited transient, rhythmic generator potentials, and altered excitability without changing the membrane potential or input resistance. These effects suggest that olfactory transduction is mediated through at least three different pathways with effects on four or more components of the membrane conductance. Polychotomous pathways such as these may be important for odor discrimination and for sharpening the "odor image" generated in the olfactory epithelium.

Action Potentials↗
PubMed1989

Multiple voltage-sensitive K+ channels regulate dendritic excitability in cerebellar Purkinje neurons.

Ionic conductances present in the dendritic region of the cerebellar Purkinje neuron were studied using the single-channel and whole-cell recording methods. Several types of voltage-sensitive K+ channels including a Ca2+ activated K+ channel were found to be a prominent components of the dendritic membrane. All patches studied contained K+ channel types and most patches contained more than one K+ channel type. In cell attached recordings, K+ channel activity was associated with the late phase of spontaneous action potentials suggesting a functional relationship. These data demonstrate that voltage-sensitive ion channels contribute to dendritic excitability and suggest that the transduction and integration of synaptic signals may involve both active and passive ionic conductances.

Adrenergic alpha-Antagonists↗
PubMed1989

Two types of nicotinic acetylcholine receptor channels at slow fibre end-plates of the garter snake.

1. Two different types of acetylcholine receptor channels can be detected on the post-junctional membrane of slow muscle fibres in garter snakes. Here they are designated T-type and S-type channels. Only T-type channels can also be found at twitch fibre neuromuscular junctions. 2. The physiological properties of slow fibre T-type channels are similar to those of acetylcholine receptor channels in end-plates of twitch fibres in these animals. 3. S-type channels had a smaller conductance than T-type channels (32 vs. 49 pS), but a similar reversal potential near 0 mV. 4. Both S- and T-type channels were found together in most patches of slow fibre end-plate membrane, but some patches displayed just one type or the other. 5. The activity of both S- and T-type channels desensitized in the presence of micromolar concentrations of acetylcholine. S-type channels desensitized less rapidly and less completely than did T-type channels. 6. Desensitized channels of both types recovered and produced bursts of activity, then went silent again. During a burst, channels did not appear to change type. 7. The activation of channels of either type was not correlated with activity in channels of the other type. 8. The open-duration distribution of S-type channels required two exponential components to be well fitted, with time constants in the range of 1-2 ms and 3-10 ms. In contrast, the open-duration distribution of T-type channels was a single exponential with a time constant similar in magnitude to the slower S-type component. 9. Desensitization-resistant S-type acetylcholine receptor channels could allow slow muscle end-plates to retain their sensitivity to acetylcholine during periods of heavy use. Under non-desensitizing conditions, differences in the decay properties of slow fibre end-plate currents compared to those in twitch fibres can be explained by the activation kinetics of S-type channels.

Acetylcholine↗
PubMed1988

Single subunits of the GABAA receptor form ion channels with properties of the native receptor.

The alpha and beta subunits of the gamma-aminobutyric acidA (GABAA) receptor were expressed individually in Xenopus oocytes by injection of RNA synthesized from their cloned DNAs. GABA-sensitive chloride channels were detected several days after injection with any one of three different alpha RNAs (alpha 1, alpha 2, and alpha 3) or with beta RNA. The channels induced by each of the alpha-subunit RNAs were indistinguishable, they had multiple conductance levels (10, 19, 28, and 42 picosiemens), and their activity was potentiated by pentobarbital and inhibited by picrotoxin. The beta channels usually expressed poorly but showed similar single channel conductance levels (10, 18, 27, and 40 picosiemens), potentiation by pentobarbital and inhibition by picrotoxin. The finding that both alpha and beta subunits, examined separately, form GABA-sensitive ion channels with permeation properties and regulatory sites characteristic of the native receptor suggests that the amino acid sequences that confer these properties are within the homologous domains shared by the subunits.

Animals↗
PubMed1988

Biophysical and pharmacological properties of cloned GABAA receptor subunits expressed in Xenopus oocytes.

Biochemical and immunological studies indicate that the GABAA receptor contains at least two types of subunit. Here we report that coexpression of two GABAA receptor subunit clones (alpha and beta) in Xenopus oocytes yields receptors with many biophysical properties of native GABAA receptors. These include ion selectivity, multiple single-channel conductance states, voltage-dependent gating and rectification, and complex desensitization kinetics. Furthermore, the receptors are competitively inhibited by bicuculline and display the expected allosteric and agonist effects of the barbiturate pentobarbital. The expressed receptors, however, appear to be activated by one molecule of GABA instead of two and fail to show potentiation by benzodiazepines. This implies that an additional factor(s) or subunit(s) is required for the reconstitution of a fully functional GABAA receptor.

Animals↗
PubMed1988

Apical localization of K+ channels in taste cells provides the basis for sour taste transduction.

Previous studies have shown that mudpuppy taste receptor cells respond to sour taste stimuli (weak acids) with depolarizing receptor potentials or action potentials that are blocked by the K+ channel blocker tetraethylammonium. Voltage-clamp recordings from isolated taste cells indicated that taste receptor cells exhibit a variety of voltage-dependent conductances and that acids reduce a voltage-dependent K+ current. Since taste stimuli are restricted to the apical surface of the intact tongue, only 1-2% of the taste receptor cell surface is exposed to chemical stimuli. Thus, modification of a K+ conductance would be an effective transduction mechanism in receptor cells only if the majority of K+ channels were located on the apical membrane. We have used a combination of "loose-patch" and whole-cell recording methods to map the distribution of voltage-sensitive K+ and Na+ channels on dissociated Necturus maculosus taste cells. We report here that the K+ conductance is approximately equal to 50-fold greater on apical membrane than on basolateral membrane, whereas the Na+ conductance is distributed evenly. The marked nonuniformity of the voltage-sensitive K+ conductance, together with the block of this conductance by sour stimuli, indicates that K+ current modulation is the mechanism of sour taste transduction.

Animals↗
PubMed1988

Developmental changes in K+-selective channel activity during differentiation of the Purkinje neuron in culture.

The cerebellar Purkinje neuron cultured from 20 d rat embryos is electrically inexcitable when immature, and acquires excitable membrane properties according to a programmed developmental sequence, thus providing a useful model for investigating mechanisms of CNS neuronal development. Using conventional patch-clamp techniques, we have characterized the the predominant classes of active K+-selective channels at a range of ages encompassing the entire developmental process from 5 to 29 d in vitro (DIV), and have shown pharmacologically that these channels are important contributors to the patterns of spontaneous activity generated by the Purkinje neurons. The 4 predominant classes of K+ channels that are active during steady-state depolarizing voltage commands are identified by unit conductances as the 27, 44, 70, and 100 pS channels, and show differences in several properties, including voltage dependence, sensitivity to tetraethylammonium chloride (TEA), mean open time, and time of appearance during development. Intracellular current-clamp recordings show that physiological maturation of the Purkinje neuron entails increases in the firing rate, the diversity of spike events that comprise spontaneous activity, and the sensitivity of spontaneous activity to disruption by the K+ channel blocker TEA. This increase in sensitivity to TEA correlates with the new expression of activity of the larger-conductance TEA-sensitive classes of K+ channel (70 and 100 pS types). These data show that developmental regulation of the activity of K+-selective channels contributes significantly to the ionic mechanisms that underlie the developmental transitions in spontaneous activity patterns in the Purkinje neuron.

Animals↗
PubMed1987

A polisher for patch pipets.

The design and construction of a novel apparatus to fire polish patch-clamp recording pipets is described. The device positions the polishing filament in the field of view of the high-power polishing objective and uses the mechanical stage of the microscope to hold the electrode, eliminating the need for a micromanipulator to hold either the filament or electrode.

Electrodes↗
PubMed1987

Preparation of isolated mouse olfactory receptor neurons.

A method is described for producing large numbers of isolated olfactory receptor neurons from adult mouse nasal epithelium. The dissociated neurons and other cell types isolated from nasal epithelium retain their morphology and can be identified visually. The neurons were judged to be intact and viable by trypan blue dye exclusion, the presence of olfactory marker protein (OMP), and a variety of electrophysiological measurements indicating the presence of substantial membrane potentials, low levels of intracellular Ca2+, and the ability to fire action potentials. The receptor neurons and other cell types produced by this method are amenable to the patch-clamp technique and to immunohistochemical studies.

Action Potentials↗
PubMed1987

Cyclic AMP regulates the proportion of functional acetylcholine receptors on chicken ciliary ganglion neurons.

Previous studies have shown that the number of functional acetylcholine receptors (AcChoRs) on chicken ciliary ganglion neurons in culture is considerably smaller than the total number of AcChoRs detected on the neurons by labeled receptor probes. Here we use patch-clamp recording to show that a cAMP-dependent process enhances the AcCho response of the neurons by a mechanism likely to involve an increase in the number of functional AcChoRs. The increase occurs without requiring protein synthesis and without involving a detectable increase in the total number of AcChoRs on the cell surface measured with a labeled receptor probe. The results imply that the neurons have functional and nonfunctional pools of AcChoRs and that functional receptors can be recruited from intracellular receptors or from nonfunctional receptors on the cell surface by a cAMP-dependent process. A cAMP-dependent regulation of the number of functional neurotransmitter receptors would provide a reversible mechanism by which cell-cell interactions could modulate synaptic transmission in the nervous system.

1-Methyl-3-isobutylxanthine↗
PubMed1987

Patch-clamp studies of isolated mouse olfactory receptor neurons.

Olfactory receptor neurons isolated from embryonic, neonatal, and adult mice were studied using the patch-clamp technique. Several distinct types of ion channels were characterized in patches of membrane from the neuronal soma and the dendritic knob of receptor neurons, including a 130-pS Ca++-activated K+ channel with voltage-dependent kinetics, an 80-pS Ca++-activated K+ channel with voltage-insensitive kinetics, a 25-pS K+ channel with properties similar to inward rectifiers, and a 40-pS K+ channel that was activated and then inactivated by rapid depolarization. Evidence of large-conductance (greater than 200 pS) Cl- channels, which were Ca++ insensitive and increasingly active at depolarizing membrane potentials, and voltage-activated Ca++ channels (16 pS) was also obtained. From K+ channel activity recorded from cell-attached patches, the intracellular [Ca++] was inferred to be below 0.1 microM, and the membrane potential was inferred to be approximately -50 mV. The receptor neurons had high input resistances, and action potentials could be elicited by picoampere amounts of depolarizing current. The receptor neurons responded to applied odorant molecules and to forskolin with increases in membrane conductance. These results provide a description of the membrane properties of olfactory receptor neurons and a basis for understanding their electrical activity and response to odorants.

Animals↗
PubMed1987

The properties and regulation of functional acetylcholine receptors on chick ciliary ganglion neurons.

The properties of acetylcholine receptor (AChR) channels on chick ciliary ganglion neurons in culture were examined using patch-clamp recording techniques. Acetylcholine (ACh) was applied by rapid microperfusion. Whole-cell current noise analysis revealed a single class of functional receptors on the neurons. Dose-response studies indicated a Kd of about 36 microM and a Hill coefficient of 1.5-1.7, predicting 2 ACh binding sites per receptor. Both fast and slow components of receptor desensitization were observed. Single-channel recordings from excised outside-out patches of soma membrane exposed to 2-5 microM ACh indicated a single-channel conductance of 40 pS, a reversal potential of -9 mV, a mean open duration of 1 msec, and an opening probability of 0.34. The kinetic behavior of the channels was provisionally described by a 3-closed, 1-open state model for receptor activation. In all of these properties, AChRs of ciliary ganglion neurons resemble those on skeletal muscle fibers. Growing the neurons in an elevated K+ concentration produced a 2-3-fold decrease in peak whole-cell currents induced by ACh under standard test conditions, without altering any of the single-channel properties described above. Neither changes in cholinesterase activity nor receptor distribution accounted for the decrease. Instead, calculations indicated that elevated K+ reduced the ACh response by decreasing the number of functional AChRs on the neurons. No K+-dependent decrease is observed, however, in the number of total receptors on the neurons detected either by a monoclonal antibody specific for the receptor or by an alpha-neurotoxin that binds to the receptor and blocks its function. Moreover, the number of receptors detected by the 2 probes is at least 10-fold greater than the calculated number of functional receptors. The findings suggest that only a small fraction of the AChRs on the neuronal surface is functional and that the cell can alter the ratio of functional and nonfunctional receptors in response to growth conditions.

Acetylcholine↗
PubMed1978

An analysis of the dose-response relationship at voltage-clamped frog neuromuscular junctions.

1. Frog neuromuscular junctions were viewed with Nomarski optics and voltage clamped. Agonist was applied ionophoretically and agonist concentrations were measured using a micro-electrode sensitive to quaternary amines. 2. The dose-response relationship was studied using the agonists carbamylcholine, suberyldicholine and hydroxyphenyl-propyltrimethylammonium. 3. With all of these agonists, it appeared that the ACh receptor could be active when either one or two agonist binding sites were occupied. The receptor was much more likely to be active when both sites were occupied. Agonist dissociation constants and receptor activation probabilities were estimated by non-linear regression techniques for several possible receptor activation schemes.

Animals↗
PubMed1976

Characterization of drug iontophoresis with a fast microassay technique.

The iontophoretic release of drugs from micropipettes into free (Ringer's) solution was described using an ion-selective microelectrode assay method. This characterization, with a temporal resolution of 20 ms, showed that the equilibrium rate of drug transport was not linearly proportional to release current; the departure from linearity was increased by backing current and the result was demonstrated with analytically derived drug release functions. The general relation between the drug transport rate and release current was independent of the specific drug or pipette resistance; no functional relation was observed that might quantitatively predict this dependence without prior use of the assay. The diffusion coefficients at 25 degrees C in frog Ringer's of the drugs used in this study, all neuromuscular agonists, were determined: all values X 10(6) cm2/s; acetylcholine 6.11 +/- 0.30; carbamylcholine 7.44 +/- 0.34; 3-(m-hydroxyphenyl) prophyltrimethyl ammonium 5.79 +/- 0.13.

Acetylcholine↗