Acid-sensing ion channel-1a (ASIC1a) mediates Hϩ -gated current to influence normal brain physiology and impact several models of disease. Although ASIC2 subunits are widely expressed in brain and modulate ASIC1a current, their function remains poorly understood. We identified ASIC2a in dendrites, dendritic spines, and brain synaptosomes. This localization largely relied on ASIC2a binding to PSD-95 and matched that of ASIC1a, which does not coimmunoprecipitate with PSD-95. We found that ASIC2 and ASIC1a associated in brain, and through its interaction with PSD-95, ASIC2 increased ASIC1a localization in dendritic spines.

Rationale Acid-Sensing Ion Channels (ASICs) are Na+ channels that are activated by acidic pH. Their expression in cardiac afferents and remarkable sensitivity to small pH changes has made them leading candidates to sense cardiac ischemia. Objective Four genes encode six different ASIC subunits, however it is not yet clear which of the ASIC subunits contribute to the composition of ASICs in cardiac afferents. Methods and Results Here we labeled cardiac afferents using a retrograde tracer dye in mice, which allowed for patch-clamp studies of murine cardiac afferents. We found that a higher percentage of cardiac sensory neurons from the dorsal root ganglia (DRG) respond to acidic pH and generated larger currents compared to those from the nodose ganglia (NG). The ASIC-like current properties of the cardiac DRG neurons from wild-type mice most closely matched the properties of ASIC2a/3 heteromeric channels. This was supported by studies in ASIC null mice: acid-evoked currents from ASIC3 −/− cardiac afferents matched the properties of ASIC2a channels, and currents from ASIC2 −/− cardiac afferents matched the properties of ASIC3 channels. Conclusions We conclude that ASIC2a and -3 are the major ASIC subunits in cardiac DRG neurons, and provide potential molecular targets to attenuate chest pain and deleterious reflexes associated with cardiac disease.

AcidAcid-sensing ion channels (ASICs) 3 are H ϩ -gated members of the DEG/ENaC ion channel family. In mammals, ASICs include four genes (ASIC1, -2, -3, and -4) that encode for six subunits (ASIC1 and -2 both have alternative splice transcripts as follows: ASIC1a, -1b, -2a, and -2b) (1, 2). Functional ASIC channels consist of a complex of three subunits (3), and they are principally expressed in neurons in the central nervous system and in peripheral sensory nerves. In the brain, the isoform ASIC1a is the best studied, and evidence suggests it plays a role in learning and memory. Genetic or pharmacological perturbation of ASIC1a affects spatial memory, eye-blink conditioning, and it seems to be particularly important for fear-related learning and behaviors (4 -6). ASIC1a also has important functions during pathological conditions, including stroke, seizures, depression, and brain tumors (7-10).For several reasons, ASIC channels are ideally positioned to sense changes in brain interstitium. First, the structure of ASICs is unique for ion channels in that ϳ70% of the entire protein consists of a single large extracellular loop. Second, ASIC1a homomeric channels are profoundly sensitive to subtle pH changes; the threshold of activation is ϳ7.0, and half-maximal activation occurs at pH ϳ6.8 (11), which is well within the range that occurs in the brain interstitium during ischemia, seizures, or spreading depression (12, 13). In fact, loss of ASIC1a abolishes currents in central nervous system neurons evoked by extracellular pH changes in the range between 7.2 and 6.0 (6). Third, multiple other chemicals that are released by metabolically stressed brain cells can potentiate ASICs. For example, ASIC currents are increased by physiological concentrations of lactate, ATP, or arachidonic acid (14 -16), all of which are released into the interstitium during brain ischemia (17)(18)(19).The recently resolved ASIC1a crystal structure revealed the surprising finding that three Cl Ϫ ions were bound to the channel complex in the extracellular domain (3). These sites are coordinated by two nearby residues (Arg-310 and Glu-314) on an ␣-helix of one subunit, a residue (Lys-212) from an adjacent subunit, and are almost completely conserved between all H ϩ -gated ASIC isoforms. Extracellular anions are known to modulate a wide variety of ion channels, including the ASICrelated epithelial sodium channel ENaC (20 -23). However, the significance of Cl Ϫ binding to ASIC channels is unknown. Here, we investigated the effect of extracellular Cl Ϫ and other anions on heterologously expressed ASIC1a as well as native ASICs in mammalian central nervous system neurons. EXPERIMENTAL PROCEDURESHeterologous Expression of cDNA in CHO Cells-Mouse ASIC1a was cloned as described previously (11). The mutations ASIC1a K211A , ASIC1a R309A , and ASIC1a E313A (the residues are numbered per the mouse ASIC1a sequence) were generated by site-directed mutagenesis using the QuikChange kit (Stratagene, La Jolla, CA) and sequenced at the University of Iowa DNA c...

Dynamic obstruction to left ventricular outflow in patients with hypertrophic cardiomyopathy usually occurs when the anterior mitral leaflet moves forward in systole and approaches or contacts the ventricular septum. However, we have recently identified, by M mode and two-dimensional echocardiography, 21 patients with hypertrophic cardiomyopathy who had a unique pattern of mitral valve motion characterized by abnormal mitral valve coaptation and systolic anterior motion of the posterior mitral leaflet. This abnormality of mitral valve motion was most reliably identified with twodimensional echocardiography in views of the left ventricle obtained from the apex. At end-diastole the anterior and posterior mitral leaflets did not appear to coapt at their distal free margins. Rather, at mitral valve closure, the anterior mitral leaflet contacted the basal portion of posterior mitral leaflet. Subsequently, during systole the "residual" distal portion of posterior mitral leaflet approached or contacted the ventricular septum. Morphologic observations in nine other patients with hypertrophic cardiomyopathy suggested that systolic anterior motion of the posterior mitral leaflet is due to elongation of the middle scallop of the posterior leaflet, which probably comes into apposition with the ventricular septum during systole by passing through the space created by the normal pattern of chordal attachments onto the anterior mitral leaflet. Of the 16 patients who underwent cardiac catheterization, nine had basal subaortic gradients of 20 to 85 mm Hg, which were apparently due to moderate or marked systolic anterior motion of the posterior mitral leaflet. Ventricular septal myotomy-myectomies were performed in two patients and resulted in markedly diminished systolic anterior motion of the posterior mitral leaflet in each and abolition of subaortic gradient in the one patient who underwent postoperative cardiac catheterization. Hence, in patients with hypertrophic cardiomyopathy, systolic anterior motion of the posterior mitral leaflet (1) is not uncommon (identifiable in about 10% of a consecutively studied series of patients), (2) constitutes a previously undescribed mechanism for dynamic subaortic obstruction, and (3) is due to a malformation of the posterior mitral leaflet. Circulation 68, No. 2, 282-293, 1983. A SUBSTANTIAL PROPORTION of patients with hypertrophic cardiomyopathy demonstrate obstruction to left ventricular outflow.'-5 Most available angiographic6' 7 and echocardiographic-25 data appear to indicate that this dynamic subaortic obstruction occurs when the anterior mitral leaflet (AML) (with or without its attached chordae tendineae) approaches or comes into contact with the ventricular septal endocardium. Others have suggested that the chordae tendineae or papillary muscles may primarily produce left