Acid-sensing Ionic Channels Research Articles

Properties and Differential Expression of H+ Receptors in Dorsal Root Ganglia: Is a Labeled-Line Coding for Acid Nociception Possible?

Pain by chemical irritants is one of the less well-described aspects of nociception. The acidic substance is the paradigm of the chemical noxious compound. An acidic insult on cutaneous, subcutaneous and muscle tissue results in pain sensation. Acid (or H+) has at least two main receptor channels in dorsal root ganglia (DRG) nociceptors: the heat receptor transient receptor potential vanilloid 1 (TRPV1) and the acid-sensing ionic channels (ASICs). TRPV1 is a low-sensitivity H+ receptor, whereas ASIC channels display a higher H+ sensitivity of at least one order of magnitude. In this review, we first describe the functional and structural characteristics of these and other H+-receptor candidates and the biophysics of their responses to low pH. Additionally, we compile reports of the expression of these H+-receptors (and other possible complementary proteins) within the DRG and compare these data with mRNA expression profiles from single-cell sequencing datasets for ASIC3, ASIC1, transient receptor potential Ankiryn subtype 1 (TRPA1) and TRPV1. We show that few nociceptor subpopulations (discriminated by unbiased classifications) combine acid-sensitive channels. This comparative review is presented in light of the accumulating evidence for labeled-line coding for most noxious sensory stimuli.

Expression and functions of ASIC1 in the zebrafish retina

It has been demonstrated that acid sensing ionic channels (ASICs) are present in the central and peripheral nervous system of mammals, including the retina. However, it remains unclear whether the zebrafish retina also expresses ASICs. In the present study, the expression and distribution of zasic1 were examined in the retina of zebrafish. Both zasic1 mRNA and protein expressions were detected in the adult zebrafish retina. A wide distribution of ASIC1 in zebrafish retina was confirmed using whole mount in situ hybridization and immunohistochemistry study. Acidosis-induced currents in the isolated retinal ganglion cells (RGCs) were also recorded using whole cell patch clamping. Moreover, blockade of ASICs channel significantly reduced the locomotion of larval zebrafish in response to light exposure. In sum, our data demonstrate the presence of ASIC1 and its possible functional relevance in the retina of zebrafish.

The Aminoglycosides Modulate the Acid-Sensing Ionic Channel Currents in Dorsal Root Ganglion Neurons from the Rat

Acid-sensing ionic channels (ASICs) have been shown to have a significant role in a growing number of physiological and pathological processes, such as nociception, synaptic transmission and plasticity, mechanosensation, and acidosis-induced neuronal injury. The discovery of pharmacological agents targeting ASICs has significant therapeutic potential and use as a research tool. In our work, we studied the action of transient perfusion (5-15 s) of aminoglycosides (AGs) (streptomycin and neomycin) on the proton-gated ionic currents in dorsal root ganglion (DRG) neurons of the rat and in human embryonic kidney (HEK)-293 cells. In DRG neurons, streptomycin and neomycin (30 microM) produced a significant, concentration-dependent, and reversible reduction in the amplitude of the proton-gated current, and a slowing of the desensitization rate of the ASIC current. Gentamycin (30 microM) also showed a significant reversible action on the ASIC currents. The curves of the pH effect for streptomycin and neomycin indicated that their effect was not significantly affected by pH. In HEK-293 cells, streptomycin (30 microM) produced a significant reduction in the amplitude of the proton-gated current. Neomycin and gentamycin had no significant action. Reduction of extracellular Ca(2+) concentration produced a significant increase in the action of streptomycin and neomycin on the desensitization time course of ASIC currents. These results indicate that ASICs are molecular targets for AGs, which may contribute to the understanding of their actions on excitable cells. Moreover, AGs may constitute a source to develop novel molecules with a greater affinity, specificity, and selectivity for the different ASIC subunits.

Acid-sensing ionic-channel functional expression in the vestibular endorgans

In the vestibular system, the electrical discharge of the afferent neurons has been found to be highly sensitive to external pH changes, and acid-sensing ionic-channels (ASIC) have been found to be functionally expressed in afferent neurons. No previous attempt to assay the ASIC function in vestibular afferent neurons has been done. In our work we studied the electrical discharge of the afferent neuron of the isolated inner ear of the axolotl ( Ambystoma tigrinum) to determine the participation of proton-gated currents in the postransductional information processing in the vestibular system. Microperfusion of FMRF-amide significantly increased the resting activity of the afferent neurons of the semicircular canal indicating that ASIC currents are tonically active in the resting condition. The use of ASIC antagonists, amiloride and acetylsalicylic acid (ASA), significantly reduced the vestibular-nerve discharge, corroborating the idea that the afferent neurons of the vestibular system express ASICs that are sensitive to amiloride, ASA, and to FMRF-amide. The sensitivity of the vestibular afferent-resting discharge to the microperfusion of ASIC acting agents indicates the participation of these currents in the establishment of the afferent-resting discharge.

Mechanisms of non‐steroid anti‐inflammatory drugs action on ASICs expressed in hippocampal interneurons

The inhibitory action of non-steroid anti-inflammatory drugs was investigated on acid-sensing ionic channels (ASIC) in isolated hippocampal interneurons and on recombinant ASICs expressed in Chinese hamster ovary (CHO) cells. Diclofenac and ibuprofen inhibited proton-induced currents in hippocampal interneurons (IC(50) were 622 +/- 34 muM and 3.42 +/- 0.50 mM, respectively). This non-competitive effect was fast and fully reversible for both drugs. Aspirin and salicylic acid at 500 muM were ineffective. Diclofenac and ibuprofen decreased the amplitude of proton-evoked currents and slowed the rates of current decay with a good correlation between these effects. Simultaneous application of acid solution and diclofenac was required for its inhibitory effect. Unlike amiloride, the action of diclofenac was voltage-independent and no competition between two drugs was found. Analysis of the action of diclofenac and ibuprofen on activation and desensitization of ASICs showed that diclofenac but not ibuprofen shifted the steady-state desensitization curve to more alkaline pH values. The reason for this shift was slowing down the recovery from desensitization of ASICs. Thus, diclofenac may serve as a neuroprotective agent during pathological conditions associated with acidification.

Acid-Sensing Ionic Channels in the Rat Vestibular Endorgans and Ganglia

Acid-sensing ionic channels (ASICs) are members of the epithelial Na+ channel/degenerin (ENaC/DEG) superfamily. ASICs are widely distributed in the central and peripheral nervous system. They have been implicated in synaptic transmission, pain perception, and the mechanoreception in peripheral tissues. Our objective was to characterize proton-gated currents mediated by ASICs and to determine their immunolocation in the rat vestibular periphery. Voltage clamp of cultured afferent neurons from P7 to P10 rats showed a proton-gated current with rapid activation and complete desensitization, which was carried almost exclusively by sodium ions. The current response to protons (H+) has a pH0.5 of 6.2. This current was reversibly decreased by amiloride, gadolinium, lead, acetylsalicylic acid, and enhanced by FMRFamide and zinc, and negatively modulated by raising the extracellular calcium concentration. Functional expression of the current was correlated with smaller-capacitance neurons. Acidification of the extracellular pH generated action potentials in vestibular neurons, suggesting a functional role of ASICs in their excitability. Immunoreactivity to ASIC1a and ASIC2a subunits was found in small vestibular ganglion neurons and afferent fibers that run throughout the macula utricle and crista stroma. ASIC2b, ASIC3, and ASIC4 were expressed to a lesser degree in vestibular ganglion neurons. The ASIC1b subunit was not detected in the vestibular endorgans. No acid-pH-sensitive currents or ASIC immunoreactivity was found in hair cells. Our results indicate that proton-gated current is carried through ASICs and that ionic current activated by H+ contributes to shape the vestibular afferent neurons' response to its synaptic input.

Peripherally applied neuropeptide SF is equally algogenic in wild type and ASIC3−/− mice

RFa-related peptides play a significant role in the processing of pain in the CNS of mammals. Recently it has been found that, when applied subcutaneously, these peptides elicit a powerful algogenic effect. The question arises whether this peripheral effect can be connected with the ability of RFa-related peptides to decrease the rate of desensitization of acid sensing ionic channels (ASICs) expressed in primary sensory neurons. We have addressed this question by comparing the effects of neuropeptide SF (NPSF), mammalian RFa peptide, in ASIC3−/− and wild-type C57BL/6J mice. Knockout of ASIC3 gene results in the changes in some of the behavioral parameters. However, subcutaneous injections of the NPSF into the n.saphenous innervation area result in a clearly nociceptive behavior in both strains of mice. There is no significant difference in the total time of licking of injected paw in the ASIC3−/− (194 ± 22 s) and C57BL/6J (227 ± 25 s) animals. Thus peripheral algogenic effects of NPSF cannot be explained only in terms of their action on the ASIC3 channels and involves some other, still unidentified mechanism.

The ASICs: Signaling molecules? Modulators?

Acid-sensing ionic channels (ASICs) are almost ubiquitous in the mammalian nervous system, both at the periphery and in the brain. Strong evidence for the physiological function of these molecules has come from recent knockout experiments. Now it is clear that ASICs are important for certain sensory modalities (mechanoreception and nociception) at the periphery and for learning and memory in the brain. The actual mechanisms by which the acid-gated channels serve these functions remain unclear. The question of whether tissue pH is subject to quick fluctuations of a magnitude sufficient to activate ASICs is a crucial point that will determine the functional significance of these channels.

The Phe-Met-Arg-Phe-amide-activated Sodium Channel Is a Tetramer

The Helix aspersa Phe-Met-Arg-Phe-amide (FMRFamide)-gated sodium channel is formed by homomultimerization of several FMRFamide-activated Na+ channel (FaNaCh) proteins. FaNaCh is homologous to the subunits that compose the amiloride-sensitive epithelial sodium channel, to Caenorhabditis elegans degenerins, and to acid-sensing ionic channels. FaNaCh properties were analyzed in stably transfected human embryonic kidney cells (HEK-293). The channel was functional with an EC50 for FMRFamide of 1 microM and an IC50 (25 degreesC) for amiloride of 6.5 microM as assessed by 22Na+ uptake measurements. The channel activity was associated with the presence of a protein at the cell surface with an apparent molecular mass of 82 kDa. The 82-kDa form was derived from an incompletely glycosylated form of 74 kDa found in the endoplasmic reticulum. Formation of covalent bonds between subunits of the same complex were observed either after formation of intersubunit disulfide bonds following cell homogenization and solubilization with Triton X-100 or after use of bifunctional cross-linkers. This resulted in the formation of covalent multimers that contained up to four subunits. Hydrodynamic properties of the solubilized FaNaCh complex also indicated a maximal stoichiometry of four subunits per complex. It is likely that epithelial Na+ channels, acid-sensing ionic channels, degenerins, and the other proteins belonging to the same ion channel superfamily also associate within tetrameric complexes.

DGNaC1, a Gonad-specific Amiloride-sensitive Na+Channel

Amiloride-sensitive sodium channels have been implicated in reproductive and early developmental processes of several species. These include the fast block of polyspermy in Xenopus oocytes that follows the sperm binding to the egg or blastocoel expansion in mammalian embryo. We have now identified a gene called dGNaC1 that is specifically expressed in the gonads and early embryo in Drosophila melanogaster. The corresponding protein belongs to the superfamily of cationic channels blocked by amiloride that includes Caenorhabditis elegans degenerins, the Helix aspersa FMRF-amide ionotropic receptor (FaNaC), the mammalian epithelial Na+ channel (ENaC), and acid-sensing ionic channels (ASIC, DRASIC, and MDEG). Expression of dGNaC1 in Xenopus oocytes generates a constitutive current that does not discriminate between Na+ and Li+, but is selective for Na+ over K+. This current is blocked by amiloride (IC50 = 24 microM), benzamil (IC50 = 2 microM), and ethylisopropyl amiloride (IC50 = 49 microM). These properties are clearly different from those obtained after expression of the previously cloned members of this family, including ENaC and the human alphaENaC-like subunit, deltaNaC. Interestingly, the pharmacology of dGNaC1 is not very different from that found for the Na+ channel characterized in rabbit preimplantation embryos. We postulate that this channel may participate in gametogenesis and early embryonic development in Drosophila.