Biomedical subjects

Ana E Vázquez

Publications and source records attributed to Ana E Vázquez.

6 recordsLinked to original sources

PubMed2006

Cochlear function in mice lacking the BK channel alpha, beta1, or beta4 subunits.

Large conductance voltage- and calcium-activated potassium (BK) channels are important for regulating many essential cellular functions, from neuronal action potential shape and firing rate to smooth muscle contractility. In amphibians, reptiles, and birds, BK channels mediate the intrinsic frequency tuning of the cochlear hair cell by an electrical resonance mechanism. In contrast, inner hair cells of the mammalian cochlea are extrinsically tuned by accessory structures of the cochlea. Nevertheless, BK channels are present in inner hair cells and encode a fast activating outward current. To understand the role of the BK channel alpha and beta subunits in mammalian inner hair cells, we analyzed the morphology, physiology, and function of these cells from mice lacking the BK channel alpha (Slo-/-) and also the beta1 and beta4 subunits (beta1/4-/-). Beta1/4-/- mice showed normal subcellular localization, developmental acquisition, and expression of BK channels. Beta1/4-/- mice showed normal cochlear function as indicated by normal auditory brainstem responses and distortion product otoacoustic emissions. Slo-/- mice also showed normal cochlear function despite the absence of the BKalpha subunit and the absence of fast activating outward current from the inner hair cells. Moreover, microarray analyses revealed no compensatory changes in transcripts encoding ion channels or transporters in the cochlea from Slo-/- mice. Slo-/- mice did, however, show increased resistance to noise-induced hearing loss. These findings reveal the fundamentally different contribution of BK channels to nonmammalian and mammalian hearing and suggest that BK channels should be considered a target in the prevention of noise-induced hearing loss.

Animals↗

PubMed2005

Differential expression of small-conductance Ca2+-activated K+ channels SK1, SK2, and SK3 in mouse atrial and ventricular myocytes.

Small-conductance Ca2+-activated K+ channels (SK channels, KCa channels) have been reported in excitable cells, where they aid in integrating changes in intracellular Ca2+ with membrane potential. We recently reported for the first time the functional existence of SK2 (KCa2.2) channels in human and mouse cardiac myocytes. Here, we report cloning of SK1 (KCa2.1) and SK3 (KCa2.3) channels from mouse atria and ventricles using RT-PCR. Full-length transcripts and their variants were detected for both SK1 and SK3 channels. Variants of mouse SK1 channel (mSK1) differ mainly in the COOH-terminal structure, affecting a portion of the sixth transmembrane segment (S6) and the calmodulin binding domain (CaMBD). Mouse SK3 channel (mSK3) differs not only in the number of polyglutamine repeats in the NH2 terminus but also in the intervening sequences between the polyglutamine repeats. Full-length cardiac mSK1 and mSK3 show 99 and 91% nucleotide identity with those of mouse colon SK1 and SK3, respectively. Quantification of SK1, SK2, and SK3 transcripts between atria and ventricles was performed using real-time quantitative RT-PCR from single, isolated cardiomyocytes. SK1 transcript was found to be more abundant in atria compared with ventricles, similar to the previously reported finding for SK2 channel. In contrast, SK3 showed similar levels of expression in atria and ventricles. Together, our data are the first to indicate the presence of the three different isoforms of SK channels in heart and the differential expression of SK1 and SK2 in mouse atria and ventricles. Because of the marked differential expression of SK channel isoforms in heart, specific ligands for Ca2+-activated K+ currents may offer a unique therapeutic opportunity to modify atrial cells without interfering with ventricular myocytes.

Amino Acid Sequence↗

PubMed2004

Evaluating cochlear function and the effects of noise exposure in the B6.CAST+Ahl mouse with distortion product otoacoustic emissions.

Cochlear function and susceptibility to noise over-exposure were examined in the congenic mouse strain B6.CAST+Ahl (B6.CAST) and compared to these same features in the CAST/Ei (CAST) and C57BL/6J (C57) parental strains. For both types of comparisons, the primary measure was the distortion-product otoacoustic emissions (DPOAE) at 2f1-2f2. Our assumption was that the B6.CAST mouse was corrected for the early onset age-related hearing loss (AHL) exhibited by one of its parental strains (C57) by the age-resistant properties of its other parental strain (CAST), and thus would exhibit neither AHL nor susceptibility to noise overstimulation effects. With respect to cochlear function, for 2.5-month mice, there was a tendency for DPOAEs to be slightly lower for mid-frequency primary tones for both C57 and B6.CAST mice, while the former mice showed clear AHL effects at the highest test frequency. However, by 5 months of age, the B6.CAST mice, like the CAST mice, displayed robust DPOAE levels that were significantly larger than DPOAE levels for the C57 mice, which were essentially absent for frequencies above about 30 kHz. To investigate the role of the Ahl gene in the susceptibility of the cochlea to the effects of noise over-exposure, two distinct paradigms consisting of temporary (TTS: 1-min, 105-dB SPL, 10-kHz pure tone) and permanent (PTS: 1-h, 105-dB SPL, 10-kHz octave band noise) threshold-shift protocols were used. The brief TTS exposure produced reversible reductions in DPOAEs that for both the B6.CAST and CAST mice recovered to within a few dB of their baseline levels by 3 min post-exposure. In contrast, the C57 mice recovered somewhat slower and, by 5 min post-exposure, emission levels were still 5 dB or more below their corresponding pre-exposure values. At 3 months of age, the TTS mice along with another group of naïve subjects representing the same three mouse strains were exposed to the PTS paradigm. By 4 days post-exposure, for B6.CAST and CAST mice, DPOAE levels had recovered to their pre-exposure control levels. However, DPOAEs for the C57 mice at most of the measurable frequencies were at least 10-30 dB lower than their counterpart baseline levels. Together these data suggest that the Ahl allele in the C57 strain contributes to both the early onset AHL exhibited by these mice as well as their susceptibility to both TTS and PTS over-exposures.

Animals↗

PubMed2003

Cloning and expression of a small-conductance Ca(2+)-activated K+ channel from the mouse cochlea: coexpression with alpha9/alpha10 acetylcholine receptors.

Functional interactions between ligand-gated, voltage-, and Ca(2+)-activated ion channels are essential to the properties of excitable cells and thus to the working of the nervous system. The outer hair cells in the mammalian cochlea receive efferent inputs from the brain stem through cholinergic nerve fibers that form synapses at their base. The acetylcholine released from these efferent fibers activates fast inhibitory postsynaptic currents mediated, to some extent, by small-conductance Ca(2+)-activated K+ channels (SK) that had not been cloned. Here we report the cloning, characterization, and expression of a complete SK2 cDNA from the mouse cochlea. The cDNAs of the mouse cochlea alpha9 and alpha10 acetylcholine receptors were also obtained, sequenced, and coexpressed with the SK2 channels. Human cultured cell lines transfected with SK2 yielded Ca(2+)-sensitive K+ current that was blocked by dequalinium chloride and apamin, known blockers of SK channels. Xenopus oocytes injected with SK2 in vitro transcribed RNA, under conditions where only outward K+ currents could be recorded, expressed an outward current that was sensitive to EGTA, dequalinium chloride, and apamin. In HEK-293 cells cotransfected with cochlear SK2 plus alpha9/alpha10 receptors, acetylcholine induced an inward current followed by a robust outward current. The results indicate that SK2 and the alpha9/alpha10 acetylcholine receptors are sufficient to partly recapitulate the native hair cell efferent synaptic response.

Acetylcholine↗

PubMed2003

Molecular identification and functional roles of a Ca(2+)-activated K+ channel in human and mouse hearts.

The repolarization phase of cardiac action potential is prone to aberrant excitation that is common in cardiac patients. Here, we demonstrate that this phase is markedly sensitive to Ca2+ because of the surprising existence of a Ca2+-activated K+ currents in cardiac cells. The current was revealed using recording conditions that preserved endogenous Ca2+ buffers. The Ca2+-activated K+ current is expressed differentially in atria compared with ventricles. Because of the significant contribution of the current toward membrane repolarization in cardiac myocytes, alterations of the current magnitude precipitate abnormal action potential profiles. We confirmed the presence of a small conductance Ca2+-activated K+ channel subtype (SK2) in human and mouse cardiac myocytes using Western blot analysis and reverse transcription-polymerase chain reaction and have cloned SK2 channels from human atria, mouse atria, and ventricles. Because of the marked differential expression of SK2 channels in the heart, specific ligands for Ca2+-activated K+ currents may offer a unique therapeutic opportunity to modify atrial cells without interfering with ventricular myocytes.

Amino Acid Sequence↗

PubMed2002

Gene-targeted deletion of neurofibromin enhances the expression of a transient outward K+ current in Schwann cells: a protein kinase A-mediated mechanism.

Mutations in the neurofibromatosis type 1 gene predispose patients to develop benign peripheral nerve tumors (neurofibromas) containing Schwann cells (SCs). SCs from neurofibromatosis type-1 gene (Nf1) null mutant mice showed increased levels of Ras-GTP and cAMP. The proliferation and differentiation of SCs are regulated by Ras-GTP and cAMP-mediated signaling, which have been linked to expression of K+ channels. We investigated the differential expression of K+ currents in Nf1 null mutant SCs (Nf1-/-) and their wild-type (Nf1+/+) counterparts and determined the mechanisms underlying the differences. The current densities of the sustained component of K+ currents were similar in the two genotypes. However, Nf1-/- SCs showed a significant increase (approximately 1.5-fold) in a 4-aminopyridine-sensitive transient outward K+ current (I(A)). Nonstationary fluctuation analysis revealed a significant increase in the number of functional channels in the null mutant cells. When the involvement of the Ras pathway in the modulation of the K+ current was examined using adenoviral-mediated gene transfer of a dominant-negative H-Ras N17 or the known H-Ras inhibitor (L-739,749), an additional increase in I(A) was observed. In contrast, protein kinase A (PKA) inhibitors, H89 and [PKI(2-22)amide] attenuated the enhancement of the current in the Nf1-/- cells, suggesting that the increase in I(A) was mediated via activation of protein kinase A. The unitary conductance of the channel underlying I(A) was unaltered by inhibitors of PKA. Activation of I(A) is thus negatively regulated by Ras-GTP and positively regulated by PKA.

Animals↗