Biomedical subjects

B W Ache

Publications and source records attributed to B W Ache.

At least 19 recordsLinked to original sources

PubMed2006

Intrinsically bursting olfactory receptor neurons.

Rhythmically bursting neurons are fundamental to neuronal network function but typically are not considered in the context of primary sensory signaling. We now report intrinsically bursting lobster primary olfactory receptor neurons that respond to odors with a phase-dependent burst of action potentials. Rhythmic odor input as might be generated by sniffing entrains the intrinsic bursting rhythm in a concentration-dependent manner and presumably synchronizes the ensemble of bursting cells. We suggest such intrinsically bursting olfactory receptor cells provide a novel way for encoding odor information.

Action Potentials↗

PubMed2006

Block by amiloride derivatives of odor-evoked discharge in lobster olfactory receptor neurons through action on a presumptive TRP channel.

Amiloride and its derivatives inhibit a number of sensory transduction processes, including some types of chemosensory transduction. Here, we report that pyrazine derivatives of amiloride reversibly inhibit odorant-evoked activity in lobster olfactory receptor neurons. The potency sequence is as follows-(IC50, mM): 5-(N,N-hexamethylene)amiloride (0.015) approximately 5-(N-methyl-N-isobutyl)amiloride (0.02) approximately 5-(N-ethyl-N-isopropyl)amiloride (0.03) > 5-(N,N-dimethyl)amiloride (0.48); 3',4'-dichlorobenzamil (0.4), phenamil (0.5), and amiloride itself (2) are ineffective. The same derivatives with the similar potency sequence also block a presumptive transient receptor potential (TRP) channel that is the likely downstream target of phosphoinositide signaling in these cells. Our results suggest that pyrazine derivatives of amiloride are useful probes to study more detailed mechanisms of chemosensory transduction in this system and possibly in other chemosensory systems in which TRP channels are the known or suspected downstream effector.

Amiloride↗

PubMed2004

Modulation of the olfactory CNG channel by Ptdlns(3,4,5)P3.

Recent data suggest that the 3-phosphoinositides can modulate cyclic nucleotide signaling in rat olfactory receptor neurons (ORNs). Given the ability of diverse lipids to modulate ion channels, we asked whether phosphatidylinositol 3,4,5-trisphosphate (PIP3) can regulate the olfactory cyclic nucleotide-gated (CNG) channel as a possible mechanism for this modulation. We show that applying PIP3 to the intracellular side of inside-out patches from rat ORNs inhibits activation of the olfactory CNG channel by cAMP. The effect of PIP3 is immediate and partially reversible, and reflects an increase in the EC50 of cAMP, not a reduction in the single-channel current amplitude. The effect of PIP3 is significantly stronger than that of PIP2; other phospholipids tested have no appreciable effect on channel activity. PIP3 similarly inhibits the recombinant heteromeric (A2/A4) and homomeric (A2) olfactory CNG channel expressed in HEK293 cells, suggesting that PIP3 acts directly on the channel. These findings indicate that 3-phosphoinositides can be functionally important regulators of CNG channels.

Animals↗

PubMed2001

An odorant-suppressed Cl- conductance in lobster olfactory receptor cells.

Odorants evoke an outward current in cultured lobster olfactory receptor neurons voltage clamped at -60 mV. The reversal potential of the outward current is independent of the reversal potential of potassium, but shifts with imposed changes in the reversal potential of chloride. The slope of the current-voltage relationship is negative, suggesting that the current is mediated by the odorant suppressing a steady-state conductance. Anthracene-9-carboxylic acid, a specific chloride channel blocker, reversibly inhibits the steady-state conductance. Local application of odorants to the outer dendrites evokes a hyperpolarizing receptor potential in lobster olfactory receptor neurons current-clamped at -70 mV in situ. Consistent with the current characterized in the cultured cells, hyperpolarizing receptor potentials in some cells are voltage sensitive, blocked by anthracene-9-carboxylic acid and associated with a decrease in membrane conductance. These results support the hypothesis that odorants suppress a steady-state chloride conductance in lobster olfactory receptor neurons. Evidence that the chloride conductance can coexist with a 4-aminopyridine-blockable potassium conductance reported earlier in these cells suggests that two distinct mechanisms can mediate odorant-evoked inhibition in lobster olfactory receptor neurons.

Action Potentials↗

PubMed2001

Odor-stimulated phosphatidylinositol 3-kinase in lobster olfactory receptor cells.

Two antagonists of phosphoinositide 3-OH kinases (PI3Ks), LY294002 and Wortmannin, reduced the magnitude of the receptor potential in lobster olfactory receptor neurons (ORNs) recorded by patch clamping the cells in vivo. An antibody directed against the c-terminus of human PI3K-P110 beta detected a molecule of predicted size in the outer dendrites of the ORNs. Two 3-phosphoinositides, PI(3,4)P(2) (1--4 microM) and PI(3,4,5)P(3) (1--4 microM) applied to the cytoplasmic side of inside-out patches taken from cultured lobster ORNs, reversibly activated a Na(+)-gated channel previously implicated in the transduction cascade in these cells. 3-Phosphoinositides were the most effective phosphoinositide (1 microM) in enhancing the open probability of the channel. Collectively, these results implicate 3-phosphoinositides in lobster olfactory transduction and raise the need to consider the 3-phosphoinositide pathway in olfactory transduction.

Androstadienes↗

PubMed2000

Characterization of a phosphoinositide-mediated odor transduction pathway reveals plasma membrane localization of an inositol 1,4, 5-trisphosphate receptor in lobster olfactory receptor neurons.

The role of phosphoinositide signaling in olfactory transduction is still being resolved. Compelling functional evidence for the transduction of odor signals via phosphoinositide pathways in olfactory transduction comes from invertebrate olfactory systems, in particular lobster olfactory receptor neurons. We now provide molecular evidence for two components of the phosphoinositide signaling pathway in lobster olfactory receptor neurons, a G protein alpha subunit of the G(q) family and an inositol 1,4, 5-trisphosphate-gated channel or an inositol 1,4,5-trisphosphate (IP(3)) receptor. Both proteins localize to the site of olfactory transduction, the outer dendrite of the olfactory receptor neurons. Furthermore, the IP(3) receptor localizes to membranes in the ciliary transduction compartment of these cells at both the light microscopic and electron microscopic levels. Given the absence of intracellular organelles in the sub-micron diameter olfactory cilia, this finding indicates that the IP(3) receptor is associated with the plasma membrane and provides the first definitive evidence for plasma membrane localization of an IP(3)R in neurons. The association of the IP(3) receptor with the plasma membrane may be a novel mechanism for regulating intracellular cations in restricted cellular compartments of neurons.

Amino Acid Sequence↗

PubMed1999

Effects of phosphatidylinositol 4,5-bisphosphate and phosphatidylinositol 4-phosphate on a Na+-gated nonselective cation channel.

Olfactory receptor neurons in the lobster express a nonselective cation channel that is activated by intracellular Na+ and carries a substantial part of the depolarizing receptor current. Here, we show that phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] and phosphatidylinositol 4-phosphate [PI(4)P] applied to the intracellular face of cell-free patches activate the channel in the absence of Na+ and that antibodies against the respective phospholipids irreversibly inhibit the evoked activity. Further, we show that applying PI(4,5)P2 or PI(4)P in the presence of Na+ decreases the concentration of Na+ required to activate the channel from an EC50 of 74 to 22 mM for PI(4,5)P2 and to 29 mM for PI(4)P, respectively. Na+-gated channel activity was irreversibly inhibited by monoclonal antibodies against PI(4,5)P2 and PI(4)P in patches never exposed to exogenous phosphatidylinositols, suggesting that endogenous inositol phospholipids are required for the activation of the channel by intracellular Na+. Our findings suggest that PI(4,5)P2 and/or PI(4)P may serve as intracellular signaling molecules in these primary sensory neurons and provide a general mechanism to explain how the sensitivity of Na+-gated channels to Na+ could be much greater in intact cells than in excised membrane patches.

Animals↗

PubMed1999

Calcium regulation of cyclic nucleotide signaling in lobster olfactory receptor neurons.

An elevated free Ca2+ concentration reduces odor-stimulated production of cyclic AMP (cAMP) in the outer dendritic membranes of lobster olfactory receptor neurons in vitro. This effect can occur within 50 ms of odor stimulation. The effect is concentration-dependent at submicromolar concentrations of free Ca2+. An elevated free Ca2+ concentration also reduces basal and forskolin-stimulated cAMP levels in a concentration-dependent manner, suggesting that Ca2+ is not targeting the activation of the odor receptor/G protein complex. The degradation of synthetic cAMP by phosphodiesterases is not enhanced by an increased free Ca2+ concentration, suggesting that Ca2+ acts by down-regulating the olfactory adenylyl cyclase. Western blot analysis of the lobster olfactory sensilla that contain the outer dendrites reveals a protein in the transduction zone with a molecular mass of approximately 138 kDa that is immunoreactive to an antiserum against adenylyl cyclase type III. Given earlier evidence that Ca2+ potentially enters the receptor cell through odor-activated inositol 1,4,5-trisphosphate-gated channels, our results suggest a possible route for cross talk between the cyclic nucleotide and the inositol phospholipid signaling pathways in lobster olfactory receptor neurons.

1-Methyl-3-isobutylxanthine↗

PubMed1998

Organizational complexity in lobster olfactory receptor cells.

The current working model of transduction in lobster olfactory receptor cells suggests that: (1) inositol-1,4,5-triphosphate (IP3) is the excitatory olfactory second messenger in these cells; (2) activation of the cell also involves a secondary, current-carrying channel; and (3) the phosphoinositol pathway works in parallel to a second, cyclic nucleotide-mediated signaling pathway that provides input of opposite polarity into the cell. The complexity of intracellular signaling in lobster olfactory receptor cells renders the cells capable of fine tuning, and even integrating, the signal they send to the brain.

Animals↗

PubMed1998

Sodium-gated cation channel implicated in the activation of lobster olfactory receptor neurons.

The role of Na+-activated channels in cellular function, if any, is still elusive. We have attempted to implicate a Na+-activated nonselective cation channel in the activation of lobster olfactory receptor neurons. We show that a Na+-activated channel occurs in the odor-detecting outer dendrites. With the use of pharmacological blockers of the channel together with ion substitution, we show that a substantial part of the odor-evoked depolarization in these cells can be ascribed to a Na+-activated conductance. We hypothesize, therefore, that the Na+-activated channel amplifies the receptor current as a result of being secondarily activated by the primary odor transduction pathway.

Animals↗

PubMed1998

Na+-gated nonselective cation channel from lobster olfactory projection neurons.

Na+-gated nonselective cation channel from lobster olfactory projection neurons. J. Neurophysiol. 80: 3387-3391, 1998. A nonselective cation channel specifically activated by intracellular Na+ was identified in cell-free patches taken from cultured lobster olfactory projection neurons. Na+ reversibly activates the channel in a concentration-dependent manner, with a "half-effect" Na+ concentration of 76.4 mM at -60 mV. The conductance of the channel is 32 pS. The channel is permeable to both alkali metal (Li+ > Na+ > K+ > Rb+ > Cs+) and divalent (Ca2+ > Mn2+ > Sr2+ > Mg2+ > Ba2+ > Na+) cations. The presence of a channel with the ability to generate plateau potentials suggests that the channel may potentially contribute to oscillatory behavior in these olfactory interneurons.

Animals↗

PubMed1997

Gating and conduction properties of a sodium-activated cation channel from lobster olfactory receptor neurons.

The gating and conduction properties of a channel activated by intracellular Na+ were studied by recording unitary currents in inside-out patches excised from lobster olfactory receptor neurons. Channel openings to a single conductance level of 104 pS occurred in bursts. The open probability of the channel increased with increasing concentrations of Na+. At 210 mm Na+, membrane depolarization increased the open probability e-fold per 36.6 mV. The distribution of channel open times could be fit by a single exponential with a time constant of 4.09 msec at -60 mV and 90 mm Na+. The open time constant was not affected by the concentration of Na+, but was increased by membrane depolarization. At 180 mm Na+ and -60 mV, the distribution of channel closed times could be fit by the sum of four exponentials with time constants of 0.20, 1.46, 8.92 and 69.9 msec, respectively. The three longer time constants decreased, while the shortest time constant did not vary with the concentration of Na+. Membrane depolarization decreased all four closed time constants. Burst duration was unaffected by the concentration of Na+, but was increased by membrane depolarization. Permeability for monovalent cations relative to that of Na+ (PX/PNa), calculated from the reversal potential, was: Li+ (1.11) > Na+ (1.0) > K+ (0.54) > Rb+ (0.36) > Cs+ (0.20). Extracellular divalent cations (10 mm) blocked the inward Na+ current at -60 mV according to the following sequence: Mn2+ > Ca2+ > Sr2+ > Mg2+ > Ba2+. Relative permeabilities for divalent cations (PY/PNa) were Ca2+ (39.0) > Mg2+ (34.1) > Mn2+ (15.5) > Ba2+ (13.8) > Na+ (1.0). Both the reversal potential and the conductance determined in divalent cation-free mixtures of Na+ and Cs+ or Li+ were monotonic functions of the mole fraction, suggesting that the channel is a single-ion pore that behaves as a multi-ion pore when the current is carried exclusively by divalent cations. The properties of the channel are consistent with the channel playing a role in odor activation of these primary receptor neurons.

Animals↗

PubMed1997

Immunocytochemical analysis of glomerular regionalization and neuronal diversity in the olfactory deutocerebrum of the spiny lobster.

Antibodies against serotonin, dopamine, FMRF amide, substance P, and molluscan small cardioactive peptide (SCPB) were used to differentiate glomeruli and neurons in the olfactory deutocerebrum of the spiny lobster, Panulirus argus. Immunoreactivity to these antibodies identified distinct regions within individual columnar glomeruli of the olfactory lobe (OL), but not within the spherical glomeruli of the accessory lobe (AL). Glomeruli in the lateral, central, and medial layers of the AL, however, had different patterns of immunoreactivity. The immunostainings differentiated six types of local interneurons and three types of centrifugal projection neurons. Local interneurons included: (1) a "dorsal giant" neuron with serotonin- and FMRF amide-like immunoreactivity arborizing in most or all glomeruli of the OL, in the glomeruli of the medial and lateral layer of the AL and in the unstructured olfactory globular tract neuropil, (2) three large OL "core" neurons, two with serotonin-like and one with FMRF amide-like immunoreactivity innervating many OL glomeruli, (3) several hundred small, globuli-type OL "core" neurons with serotonin- and FMRF amide-like immunoreactivity, (4) thousands of small, globuli-type neurons with FMRF amide- and/or substance P-like immunoreactivity connecting the OL with the central layer of the AL, (5) thousands of small, globuli-type AL interneurons with substance P like immunoreactivity and additional arborizations in the unstructured deutocerebral tract neuropil, and (6) many small, globuli-type OL "rim" neurons with FMRF amide- and/or SCPB-like immunoreactivity. Centrifugal projection neurons included two that targeted the soma clusters and a pair of large neurons with dopamine-like immunoreactivity that originated in the lateral protocerebrum and arborized in the OL and AL glomeruli. Only few ascending projection neurons and no olfactory afferents were labeled. These results suggest that in the spiny lobster neurochemically distinct subpopulations of local interneurons constitute functionally distinct regions within individual OL glomeruli and across groups of AL glomeruli.

Animals↗

PubMed1997

Ionotropic GABA receptor from lobster olfactory projection neurons.

This study reports an ionotropic GABA (gamma-aminobutyric acid) receptor in projection neurons acutely dissociated from the olfactory lobe of the brain of the spiny lobster and analyzed by whole cell and cell-free patch-clamp recording. GABA evokes a macroscopic current in the cells that is linear from -100 to + 100 mV, reverses at the imposed chloride equilibrium potential, has a permeability sequence of Cl- > acetate > bicarbonate > phosphate > propionate and SCN- > Br- > I- > Cl- > F-, and is reversibly blocked by the Cl channel blocker picrotoxin but not tert-butylbicyclophosphorothionate (TBPS). The current is bicuculline insensitive and activated by muscimol, isoguvacine, cis-4-aminocrotonic acid (CACA), and trans-aminocrotonic acid (TACA), as well as by the GABA(C)-receptor antagonists 4,5,6,7-tetrahydroisoxazolo [5,4,-c]pyridin-3-ol (THIP), 3-amino-1-propanesulfonic acid (3-APS), and imidazole-4-acetic acid (I-4AA), but not the GABA(B)-receptor agonists baclofen and 3-aminopropylphosphonic acid (3-APA). Agonist potency for the receptor is TACA > muscimol > GABA > I-4AA > isoguvacine > 3-APS > CACA > THIP. Unitary chloride currents in cell-free, outside-out patches from the cells share enough of these pharmacological properties to indicate that the channel underlies the macroscopic current. The receptor mediates an inhibitory current in the cells in vivo. The receptor is similar, if not identical, to one from neurons cultured from the thoracic ganglia of the clawed lobster. The more extensive pharmacological characterization of the receptor reported here indicates that this lobster CNS receptor is pharmacologically distinct from previously characterized ionotropic GABA receptors.

Animals↗

PubMed1996

Patch-clamping arthropod olfactory receptor neurons to study mechanisms of olfactory transduction.

The olfactory organ of arthropods such as lobsters and insects consists of an array of hair-like sensilla located on the antenna. Each sensillum contains from two to several hundred primary olfactory receptor neurons. The receptor neurons can be patch-clamped in three different types of preparations: intact cells in situ, cultured cells and outer dendrites. These preparations permit using a wide range of experimental strategies to study mechanisms of olfactory transduction. The ability to integrate data from three complementary preparations is a particular advantage of using arthropod models to understand how odor information is encoded by the primary receptor cell in olfaction.

Animals↗

PubMed1995

Dual second-messenger pathways in olfactory transduction.

Increasing evidence indicates that inositol phosphate as well as cyclic nucleotide signalling pathways mediate olfactory transduction. Both pathways can target multiple ion channel effectors, suggesting that olfactory receptor cells serve as more than simple selectivity filters and that they possibly represent the first stage of olfactory integration.

Animals↗

PubMed1995

Evidence that a Gq-protein mediates excitatory odor transduction in lobster olfactory receptor neurons.

Non-hydrolysable analogs of GTP and GDP alter odor-evoked inward and outward currents in voltage-clamped cultured lobster olfactory receptor neurons. Currents of both polarities are pertussis and cholera toxin-insensitive. Antibodies directed against the alpha subunits of G(olf), G(o), G11, an internal Gq sequence, the common carboxyl terminal sequence of Gq and G11 (anti-Gq/11), and the transducin beta subunit, fail to perturb the outward current, but anti-G(o) and anti-Gq/11 selectively block the inward current. Anti-Gq/11 immunolabels a band of approximately 45 kDa by Western blot analysis, but the anti-G(o) immunolabeling is non-specific. These results suggest that the excitatory olfactory signalling pathway that leads to an odor-evoked inward current may be coupled via a member of the Gq family, while the odor-evoked outward current is transduced by a different G protein.

Amino Acid Sequence↗

PubMed1995

Na(+)-activated nonselective cation channels in primary olfactory neurons.

1. Excised inside-out patch recordings were used to describe a novel cation channel from cultured lobster olfactory receptor neurons that is activated by [Na+]i. 2. The channel is reversibly activated by intracellular Na+ as low as 5 mM. The half-effect concentration of intracellular Na+ is approximately 60 mM at -60 mV. The dependence of the channel open probability on [Na+]i is sigmoidal with a Hill coefficient of 3.1, indicating that more than one Na+ must bind to activate the channel. 3. The channel is equally permeable to Na+, K+, and Li+. In symmetrical 210 mM Na+, the open channel current-voltage relationship shows slight inward rectification at positive potentials. The slope conductance of the channel is 107 pS between -90 and 0 mV. 4. Although the channel is not activated by voltage in the absence of intracellular Na+, the gating of the channel is dependent on voltage as well as [Na+]i and [Na+]o. 5. Both intracellular Ca2+ and Mg2+ reversibly affect channel activity in a concentration-dependent manner starting at 1 microM. Ca2+ decreases both the open probability and the single channel amplitude, whereas Mg2+ decreases the open probability but has no effect on the single channel amplitude. Ba2+ (5 mM), but not 20 mM Cs+ and 100 microM amiloride, reversibly block the channel. 6. We speculate that this novel cation channel regulates neuronal excitability by accentuating the rate and/or the magnitude of depolarization of the cell to odors.

Animals↗