Biomedical subjects

A E Dubin

Publications and source records attributed to A E Dubin.

16 recordsLinked to original sources

PubMed2006

Role of peripheral hyperpolarization-activated cyclic nucleotide-modulated channel pacemaker channels in acute and chronic pain models in the rat.

Hyperpolarization-activated, cyclic nucleotide-modulated (HCN) channels contribute to rhythmic spontaneous activity in the heart and CNS. Ectopic spontaneous neuronal activity has been implicated in the development and maintenance of acute and chronic hyperalgesia, allodynia and spontaneous pain. Previously, we documented that systemic administration of ZD7288, a specific blocker of pacemaker current (I(h)), decreased ectopic activity in dorsal root ganglion (DRG) and reversed tactile allodynia in spinal nerve ligated (SNL) rats [Chaplan SR, Guo HQ, Lee DH, Luo L, Liu C, Kuei C, Velumian AA, Butler MP, Brown SM, Dubin AE (2003) Neuronal hyperpolarization-activated pacemaker channels drive neuropathic pain. J Neurosci 23:1169-1178]. Spontaneous pain is the chief clinical manifestation of peripheral nerve injury; however, a role for I(h) in spontaneous pain has not been described. Here, in further rat studies, we report that systemic administration of ZD7288 reversed spontaneous pain induced by mild thermal injury (MTI) and tactile allodynia induced by SNL and MTI. In contrast, ZD7288 did not reduce thermal hyperalgesia. An important locus of action appears to be in the skin since intraplantar (local) administration of ZD7288 completely suppressed tactile allodynia arising from MTI and SNL and reduced spontaneous pain due to MTI. Immunohistochemical staining of plantar skin sections detected HCN1-HCN4 expression in mechanosensory structures (e.g., Meissner's corpuscles and Merkel cells). Collectively, these data suggest that expression and modulation of I(h) in the peripheral nervous system, including specialized sensory structures, may play a significant role in sensory processing and contribute to spontaneous pain and tactile allodynia.

Acute Disease↗

PubMed2002

Inactivation of olfactory sensilla of a single morphological type differentially affects the response of Drosophila to odors.

The olfactory organs on the head of Drosophila, antennae and maxillary palps, contain several hundred olfactory hairs, each with one or more olfactory receptor neurons. Olfactory hairs belong to one of three main morphological types, trichoid, basiconic, and coeloconic sensilla, and show characteristic spatial distribution patterns on the surface of the antenna and maxillary palps. Here we show that targeting expression of the cell-death gene reaper to basiconic sensilla (BS) causes the specific inactivation of most olfactory sensilla of this type with no detectable effect on other types of olfactory sensilla or the structure of the antennal lobe. Our data suggest that BS are required for a normal sensitivity to many odorants with a variety of chemical structures, through a wide range of concentrations. Interestingly, however, in contrast to other odorants tested, the behavioral response of ablated flies to intermediate concentrations of propionic and butyric acids is normal, suggesting the involvement of sensilla unaffected by ectopic reaper expression, probably coeloconic sensilla that respond strongly to these two organic acids. As inactivation of BS causes an underestimation of the concentration of both acids detectable at both the highest and lowest odorants concentrations, our results suggest that concentration coding for these two odorants relies on the integration of signals from different subsets of sensilla, most likely of different morphological types.

Animals↗

PubMed2001

Pharmacological comparison of human homomeric 5-HT3A receptors versus heteromeric 5-HT3A/3B receptors.

The present study determined the detailed pharmacological profile of heterologously expressed human (h) homomeric 5-HT3A receptors in direct comparison to heteromeric h5-HT3A/3B receptors. The very minor differences in their respective pharmacological profiles indicates that the 5-HT3B receptor subunit alters, predominantly, the biophysical rather than the pharmacological properties of the 5-HT3 receptor.

Free Radical Scavengers↗

PubMed1999

The pharmacological and functional characteristics of the serotonin 5-HT(3A) receptor are specifically modified by a 5-HT(3B) receptor subunit.

While homomers containing 5-HT(3A) subunits form functional ligand-gated serotonin (5-HT) receptors in heterologous expression systems (Jackson, M. B., and Yakel, J. L. (1995) Annu. Rev. Physiol. 57, 447-468; Lambert, J. J., Peters, J. A., and Hope, A. G. (1995) in Ligand-Voltage-Gated Ion Channels (North, R., ed) pp. 177-211, CRC Press, Inc., Boca Raton, FL), it has been proposed that native receptors may exist as heteromers (Fletcher, S., and Barnes, N. M. (1998) Trends Pharmacol. Sci. 19, 212-215). We report the cloning of a subunit 5-HT(3B) with approximately 44% amino acid identity to 5-HT(3A) that specifically modified 5-HT(3A) receptor kinetics, voltage dependence, and pharmacology. Co-expression of 5-HT(3B) with 5-HT(3A) modified the duration of 5-HT(3) receptor agonist-induced responses, linearized the current-voltage relationship, increased agonist and antagonist affinity, and reduced cooperativity between subunits. Reverse transcriptase-polymerase chain reaction in situ hybridization revealed co-localization of both 5-HT(3B) and 5-HT(3A) in a population of neurons in the amygdala, telencephalon, and entorhinal cortex. Furthermore, 5-HT(3A) and 5-HT(3B) mRNAs were expressed in spleen and intestine. Our data suggest that 5-HT(3B) might contribute to tissue-specific functional changes in 5-HT(3)-mediated signaling and/or modulation.

Amygdala↗

PubMed1999

Lysophosphatidic acid stimulates neurotransmitter-like conductance changes that precede GABA and L-glutamate in early, presumptive cortical neuroblasts.

During neurogenesis in the embryonic cerebral cortex, the classical neurotransmitters GABA and L-glutamate stimulate ionic conductance changes in ventricular zone (VZ) neuroblasts. Lysophosphatidic acid (LPA) is a bioactive phospholipid producing myriad effects on cells including alterations in membrane conductances (for review, see Moolenaar et al., 1995). Developmental expression patterns of its first cloned receptor gene, lpA1/vzg-1 (Hecht et al., 1996; Fukushima et al., 1998) in the VZ suggested that functional LPA receptors were synthesized at these early times, and thus, LPA could be an earlier stimulus to VZ cells than the neurotransmitters GABA and L-glutamate. To address this possibility, primary cultures of electrically coupled, presumptive cortical neuroblast clusters were identified by age, morphology, electrophysiological profile, BrdU incorporation, and nestin immunostaining. Single cells from cortical neuroblast cell lines were also examined. Whole-cell variation of the patch-clamp technique was used to record from nestin-immunoreactive cells after stimulation by local administration of ligands. After initial plating at embryonic day 11 (E11), cells responded only to LPA but not to GABA or L-glutamate. Continued growth in culture for up to 12 hr produced more LPA-responsive cells, but also a growing population of GABA- or L-glutamate-responsive cells. Cultures from E12 embryos showed LPA as well as GABA and L-glutamate responses, with LPA-responsive cells still representing a majority. Overall, >50% of cells responded to LPA with depolarization mediated by either chloride or nonselective cation conductances. These data implicate LPA as the earliest reported extracellular stimulus of ionic conductance changes for cortical neuroblasts and provide evidence for LPA as a novel, physiological component in CNS development.

Animals↗

PubMed1998

Involvement of genes encoding a K+ channel (ether a go-go) and a Na+ channel (smellblind) in Drosophila olfaction.

We have investigated the roles of the putative cyclic nucleotide-modulated K+ channel subunit encoded by the ether a go-go (eag) gene and a voltage-gated Na+ channel, smellblind (sbl), encoded by the paralytic (para) locus in odorant responsiveness and cell excitability in Drosophila melanogaster. Three independent mutant alleles of eag revealed reduced antennal responsiveness in adult flies to a subset of odorants, all having short aliphatic side chains: ethyl butyrate (EB), propionic acid, 2-butanone and ethyl acetate (manuscript submitted). Loose patch recordings revealed that significantly fewer eag antennal neurons responded to EB compared to control neurons. As expected if Eag were involved in odor transduction, fewer EB-induced inhibitory responses were observed in eag mutants and focal application of high K+ saline to sensillae altered the excitability of the majority of neurons from wild-type, but not eag, antennae. Interestingly, there were fewer excitatory odorant responses dependent on extracellular Ca2+ in eag neurons. In contrast to the involvement of Eag in adult olfactory neuron odorant transduction, we found no evidence that adult sbl and allelic olfactory D (olfD) gene mutants were defective in their behavioral response to a complex attractive odor. Furthermore, electrophysiological analyses of adult sbl and olfD mutants revealed normal electroantennogram responses to a broad range of individual pure odorants and no changes in the excitable properties of olfactory neurons as determined by loose patch recordings.

Animals↗

PubMed1998

The K+ channel gene ether a go-go is required for the transduction of a subset of odorants in adult Drosophila melanogaster.

The functional identity of an olfactory receptor neuron is determined in part by its repertoire of responses to odorants. As an approach toward understanding the contributions of particular conductances to olfactory neuron excitability and odor discrimination, we have investigated the role of the putative cyclic nucleotide-modulated K+ channel subunit encoded by the ether a go-go (eag) gene in odorant responsiveness in Drosophila melanogaster. Four independent mutant eag alleles exhibited reduced antennal sensitivity to a subset of nine odorants, all having short aliphatic side chains: ethyl butyrate (EB), propionic acid, 2-butanone, and ethyl acetate. Significantly fewer eag antennal neurons responded to EB compared with control neurons; the proportion sensitive to 2-heptanone was similar to controls. Two aspects of the character of EB-induced excitability were affected by mutations in eag. First, fewer EB-induced inhibitory responses were observed in eag mutants, and second, fewer excitatory odorant responses dependent on extracellular Ca2+ were observed. Furthermore, modulation of neuronal excitability by membrane-permeant cyclic nucleotide analogs was largely eag dependent. Focal application of high K+ saline to sensillae altered the excitability of the majority of neurons from wild-type but not eag antennae, suggesting that Eag may have a dendritic localization.

Animals↗

PubMed1997

A cyclic nucleotide-dependent chloride conductance in olfactory receptor neurons.

Whole-cell membrane currents were recorded from olfactory receptor neurons from the neotenic salamander Necturus maculosus. Cyclic nucleotides, released intracellularly by flash photolysis of NPE-caged cAMP or NPE-caged cGMP, activated a transient chloride current. The chloride current could be elicited at constant voltage in the absence of extracellular Ca2+ as well as in the presence of 3 mM intracellular Ca2+, suggesting that the current did not require either voltage or Ca2+ transients for activation. The current could be elicited in the presence of the protein kinase inhibitors H-7 and H-89, and in the absence of intracellular ATP, indicating that activation was independent of protein kinase A activity. These results suggest that Necturus olfactory receptor neurons contain a novel chloride ion channel that may be directly gated by cyclic nucleotides.

1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine↗

PubMed1997

Voltage-activated and odor-modulated conductances in olfactory neurons of Drosophila melanogaster.

Voltage-activated currents and odor-modulated conductances were studied in cells in semi-intact Drosophila third antennal segments (the main olfactory organ) using patch-clamp techniques. All neurons expressed outward currents, and most expressed labile fast transient inward currents with kinetics similar to Na+ currents in other systems. Action potentials were detected as bipolar capacitative current transients in cell-attached or loose patches from the soma of both odor-sensitive (97%) and insensitive neurons. A mixture of odorants from five chemical classes caused an increase (approximately 70%), decrease (approximately 10%), or no effect on firing frequency in pharate adult neurons. The development of chemosensitivity was examined and odor-induced changes in action potential firing frequency were recorded in pupal antennal neurons as early as P8, a stage after completion of sensillar development. The character of odor-induced responses was more profound and complex later in development; small, tonic increases in firing frequency were observed at pupal stages P8 through P11 (ii), while in older pupae and young adults approximately 25% of the increased responses were phasic-tonic. The apical dendrite was the site of odor modulation in approximately 90% and 100% of responsive adult and early pupal neurons, respectively. Whole-cell recordings revealed that apparent nonselective cation and chloride conductances were modulated by a mixture of odorants in separate antennal neurons.

Age Factors↗

PubMed1995

TNF alpha receptor expression in rat cardiac myocytes: TNF alpha inhibition of L-type Ca2+ current and Ca2+ transients.

Tumor necrosis factor-alpha (TNF alpha) is a potentially powerful anti-neoplastic agent; however, its therapeutic usefulness is limited by its cardiotoxic and negative inotropic effects. Accordingly, studies were undertaken to gain a better understanding of the mechanisms of TNF alpha-mediated cardiodepression. Single cell RT-PCR, [125I]TNF alpha ligand binding and Western immunoblotting experiments demonstrated that rat cardiac cells predominantly express type I TNF alpha receptors (TNFRI or p60). TNF alpha inhibited cardiac L-type Ca2+ channel current (ICa) and contractile Ca2+ transients. Thus, it is possible that the negative inotropic effects of TNF alpha are the result of TNFRI-mediated blockade of cardiac excitation-contraction coupling.

Animals↗

PubMed1995

Scutoid mutation of Drosophila melanogaster specifically decreases olfactory responses to short-chain acetate esters and ketones.

A molecular-genetic approach has been taken to identify genes involved in olfactory transduction in Drosophila melanogaster. Two independent lines of research led to the finding that the dominant Scutoid (Sco) mutation causes a diminished extracellular electroantennogram response to the odorants ethyl acetate (EtAC) and acetone (AC). Sco flies showed about 4- and 2.5-fold reduced responses to EtAC and AC, respectively, compared to Canton-S wild-type and sibling control flies lacking the Sco mutation when electroantennogram recordings were made from the proximal anterior third antennal segment. The responses to five other odors from three different chemical classes were unaltered. The maximum response to either EtAC or AC was decreased with no change in apparent affinity. Responses to short-chain (but not long-chain) acetate esters and ketones were dramatically affected at all antennal locations tested. Only in the proximal quadrants were responses to ethyl acetoacetate also reduced. Most Sco revertants tested had a normal olfactory response; duplications of the region including no-ocelli partially suppress the Sco bristle as well as olfactory phenotypes. Sco adults had an impaired behavioral response to EtAC but not to banana or propionate. There was no effect of the mutation on larval chemosensory behavior or extracellularly recorded adult compound eye and ocellar visual responses. These findings suggest the involvement of Sco in an olfactory pathway in adults which is specific for short-chain acetate esters and ketones.

Acetates↗

PubMed1994

Regulation of Shaker K+ channel inactivation gating by the cAMP-dependent protein kinase.

In response to depolarization of the membrane potential, Shaker K+ channels undergo a series of voltage-dependent conformational changes, from resting to open conformations followed by a rapid transition into a long-lived closed conformation, the N-type inactivated state. Application of phosphatases to the cytoplasmic side of Shaker channels in excised inside-out patches slows N-type inactivation gating. Subsequent application of the purified catalytic subunit of the cAMP-dependent protein kinase (PKA) and ATP reverses the effect, accelerating N-type inactivation back to its initial rapid rate. Macroscopic and single-channel experiments indicate that N-type inactivation is selectively modulated. There was little or no effect on the voltage dependence and kinetics of activation. Comparison of site-directed mutant channels shows that a C-terminal consensus site for PKA phosphorylation is responsible for the modulation. Since a cell's integrative characteristics can be determined by the rate of inactivation of its voltage-dependent channels, modulation of these rates by phosphorylation is likely to have functional consequences.

Adenosine Triphosphate↗

PubMed1994

Action potentials and chemosensitive conductances in the dendrites of olfactory neurons suggest new features for odor transduction.

Odors affect the excitability of an olfactory neuron by altering membrane conductances at the ciliated end of a single, long dendrite. One mechanism to increase the sensitivity of olfactory neurons to odorants would be for their dendrites to support action potentials. We show for the first time that isolated olfactory dendrites from the mudpuppy Necturus maculosus contain a high density of voltage-activated Na+ channels and produce Na-dependent action potentials in response to depolarizing current pulses. Furthermore, all required steps in the transduction process beginning with odor detection and culminating with action potential initiation occur in the ciliated dendrite. We have previously shown that odors can modulate Cl- and K+ conductances in intact olfactory neurons, producing both excitation and inhibition. Here we show that both conductances are also present in the isolated, ciliated dendrite near the site of odor binding, that they are modulated by odors, and that they affect neuronal excitability. Voltage-activated Cl- currents blocked by 4,4'-diisothiocyanatostilbene-2,2' disulfonic acid and niflumic acid were found at greater than five times higher average density in the ciliated dendrite than in the soma, whereas voltage-activated K+ currents inhibited by intracellular Cs+ were distributed on average more uniformly throughout the cell. When ciliated, chemosensitive dendrites were stimulated with the odorant taurine, the responses were similar to those seen in intact cells: Cl- currents were increased in some dendrites, whereas in others Cl- or K+ currents were decreased, and responses washed out during whole-cell recording. The Cl- equilibrium potential for intact neurons bathed in physiological saline was found to be -45 mV using an on-cell voltage-ramp protocol and delayed application of channel blockers. We postulate that transduction of some odors is caused by second messenger-mediated modulation of the resting membrane conductance (as opposed to a specialized generator conductance) in the cilia or apical region of the dendrite, and show how this could alter the firing frequency of olfactory neurons.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗

PubMed1994

Transduction diversity in olfaction.

Odors are powerful stimuli that can focus the attention, elicit behaviors (or misbehaviors) and even resurrect forgotten memories. These actions are directed by the central nervous system, but they depend upon the initial transduction of chemical signals by olfactory receptor neurons. Electrophysiological recordings suggest that the responses of olfactory receptor neurons to odors are more diverse than was initially believed, being mediated by effects on several different conductances. Both excitatory and inhibitory responses are produced by these effects and some, if not all, odors can affect more than one component of the membrane conductance. The extent of this diversity is reviewed here, and its impact on our understanding of odor discrimination is discussed.

Animals↗

PubMed1993

Modulation of Cl-, K+, and nonselective cation conductances by taurine in olfactory receptor neurons of the mudpuppy Necturus maculosus.

Odors are transduced by processes that modulate the membrane conductance of olfactory receptor neurons. Olfactory neurons from the aquatic salamander, Necturus maculosus, were acutely isolated without enzymes and studied with a resistive whole-cell method to minimize loss of soluble intracellular constituents. 55 of 224 neurons responded to the test compound taurine at concentrations between 10 nM and 100 microM. Four different conductance changes were elicited by taurine: an increased Cl- conductance (33%), an increased nonselective cation conductance (15%), a decreased Cl- conductance (15%), and a decreased K+ conductance (15%); in addition, responses too small to be characterized were elicited in some neurons. In most cases, taurine appeared to modulate only a single conductance in any particular cell. Modulation of each conductance was dose dependent, and each response ran down quickly in the normal whole-cell mode, presumably due to washout of a diffusible component in the transduction pathway. Modulation of taurine-sensitive conductances caused either inhibitory or excitatory responses. A similar diversity of responses in vivo would produce a complex pattern of electrical activity that could encode the identity and characteristics of an odor.

Animals↗

PubMed1992

Cyclic AMP and the nicotinic response of bovine adrenal chromaffin cells.

The effects of cAMP analogs on the nicotinic responses of bovine adrenal chromaffin cells were examined by monitoring nicotine-induced whole cell currents, [3H]norepinephrine release, and membrane conductance changes. None of the three methods revealed an increased nicotinic response after treatment with cAMP analogs. The compounds did increase [3H]norepinephrine release from the cells but the effect was not exerted at the level of nicotinic receptors. Bovine adrenal chromaffin cells differ in this respect from chick ciliary ganglion neurons which do show increased nicotinic responses after treatment with cAMP analogs.

1-Methyl-3-isobutylxanthine↗