Clarithromycin increases neuronal excitability in CA3 pyramidal neurons through a reduction in GABAergic signaling

Antibiotics are used in the treatment and prevention of bacterial infections, but effects on neuron excitability have been documented. A recent study demonstrated that clarithromycin alleviates daytime sleepiness in hypersomnia patients (Trotti LM, Saini P, Freeman AA, Bliwise DL, García PS, Jenkins A, Rye DB. J Psychopharmacol 28: 697–702, 2014). To explore the potential application of clarithromycin as a stimulant, we performed whole cell patch-clamp recordings in rat pyramidal cells from the CA3 region of hippocampus. In the presence of the antibiotic, rheobase current was reduced by 50%, F-I relationship (number of action potentials as a function of injected current) was shifted to the left, and the resting membrane potential was more depolarized. Clarithromycin-induced hyperexcitability was dose dependent; doses of 30 and 300 μM clarithromycin significantly increased the firing frequency and membrane potential compared with controls ( P = 0.003, P < 0.0001). We hypothesized that clarithromycin enhanced excitability by reducing GABAA receptor activation. Clarithromycin at 30 μM significantly reduced ( P = 0.001) the amplitude of spontaneous miniature inhibitory GABAergic currents and at 300 μM had a minor effect on action potential width. Additionally, we tested the effect of clarithromycin in an ex vivo seizure model by evaluating its effect on spontaneous local field potentials. Bath application of 300 μM clarithromycin enhanced burst frequency twofold compared with controls ( P = 0.0006). Taken together, these results suggest that blocking GABAergic signaling with clarithromycin increases cellular excitability and potentially serves as a stimulant, facilitating emergence from anesthesia or normalizing vigilance in hypersomnia and narcolepsy. However, the administration of clarithromycin should be carefully considered in patients with seizure disorders. NEW & NOTEWORTHY Clinical administration of the macrolide antibiotic clarithromycin has been associated with side effects such as mania, agitation, and delirium. Here, we investigated the adverse effects of this antibiotic on CA3 pyramidal cell excitability. Clarithromycin induces hyperexcitability in single neurons and is related to a reduction in GABAergic signaling. Our results support a potentially new application of clarithromycin as a stimulant to facilitate emergence from anesthesia or to normalize vigilance.

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Characterization of synaptically mediated fast and slow inhibitory processes in piriform cortex in an in vitro slice preparation

Journal of Neurophysiology ◽

10.1152/jn.1988.59.5.1352 ◽

1988 ◽

Vol 59 (5) ◽

pp. 1352-1376 ◽

Cited By ~ 78

Author(s):

G. F. Tseng ◽

L. B. Haberly

Keyword(s):

Membrane Potential ◽

Piriform Cortex ◽

Reversal Potential ◽

Pyramidal Cells ◽

Membrane Depolarization ◽

Resting Membrane Potential ◽

Excitatory Postsynaptic Potential ◽

Pentobarbital Sodium ◽

Postsynaptic Potential ◽

Conductance Increase

1. Intracellular recordings were obtained from anatomically verified layer II pyramidal cells in slices from rat piriform cortex cut perpendicular to the surface. 2. Responses to afferent and association fiber stimulation at resting membrane potential consisted of a depolarizing potential followed by a late hyperpolarizing potential (LHP). Membrane polarization by current injection revealed two components in the depolarizing potential: an initial excitatory postsynaptic potential (EPSP) followed at brief latency by an inhibitory postsynaptic potential (IPSP) that inverted with membrane depolarization and truncated the duration of the EPSP. 3. The early IPSP displayed the following characteristics suggesting mediation by gamma-aminobutyric acid (GABA) receptors linked to Cl- channels: associated conductance increase, sensitivity to increases in internal Cl- concentration, blockage by picrotoxin and bicuculline, and potentiation by pentobarbital sodium. The reversal potential was in the depolarizing direction with respect to resting membrane potential so that the inhibitory effect was exclusively via current shunting. 4. The LHP had an associated conductance increase and a reversal potential of -90 mV in normal bathing medium that shifted according to Nernst predictions for a K+ potential with changes in external K+ over the range 4.5-8 mM indicating mediation by the opening of K+ channels and ruling out an electrogenic pump origin. 5. Lack of effect of bath-applied 8-bromoadenosine 3',5'-cyclic monophosphate (8-Br-cAMP) or internally applied ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) on the LHP and failure of high amplitude, direct membrane depolarization to evoke a comparable potential, argue against endogenous mediation of the LHP by a Ca2+ activated K+ conductance [gK(Ca)]. However, an apparent endogenously mediated gK(Ca) with a duration much greater than the LHP was observed in a low percent of layer II pyramidal cells. Lack of effect of 8-Br-cAMP also indicates a lack of dependence of the LHP on cAMP. 6. Other characteristics of the LHP that were demonstrated include: a lack of blockage by GABAA receptor antagonists, a probable voltage sensitivity (decrease in amplitude in the depolarizing direction), and an apparent brief onset latency (less than 10 ms) when the early IPSP was blocked by picrotoxin. The LHP was unaffected by pentobarbital sodium when the early IPSP was blocked by picrotoxin. 7. Both the LHP and early IPSP were blocked by low Ca2+/high Mg2+, consistent with disynaptic mediation.(ABSTRACT TRUNCATED AT 400 WORDS)

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Enhanced NMDAR-dependent epileptiform activity is controlled by oxidizing agents in a chronic model of temporal lobe epilepsy

Journal of Neurophysiology ◽

10.1152/jn.1996.76.6.4185 ◽

1996 ◽

Vol 76 (6) ◽

pp. 4185-4189 ◽

Cited By ~ 16

Author(s):

J. C. Hirsch ◽

O. Quesada ◽

M. Esclapez ◽

H. Gozlan ◽

Y. Ben-Ari ◽

...

Keyword(s):

Synaptic Transmission ◽

Ex Vivo ◽

Epileptiform Activity ◽

Hippocampal Slices ◽

Nmda Antagonist ◽

Pyramidal Cells ◽

Resting Membrane Potential ◽

Postsynaptic Potentials ◽

Epileptiform Discharges ◽

Late Part

1. Graded N-methyl-D-aspartate receptor (NMDAR)-dependent epileptiform discharges were recorded from ex vivo hippocampal slices obtained from rats injected a week earlier with an intracerebroventricular dose of kainic acid. Intracellular recordings from pyramidal cells of the CA1 area showed that glutamate NMDAR actively participated in synaptic transmission, even at resting membrane potential. When NMDAR were pharmacologically isolated, graded burst discharges could still be evoked. 2. The oxidizing reagent 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB, 200 microM, 15 min) suppressed the late part of the epileptiform burst that did not recover after wash but could be reinstated by the reducing agent tris (2-carboxyethyl) phosphine (TCEP, 200 microM, 15 min) and again abolished with the NMDA antagonist D-2-amino-5-phosphonovaleric acid (D-APV). 3. Pharmacologically isolated NMDAR-mediated responses were decreased by DTNB (56 +/- 10%, mean +/- SD, n = 6), an effect reversed by TCEP. 4. When only the fast glutamateric synaptic component was blocked, NMDA-dependent excitatory postsynaptic potentials (EPSPs) could be evoked despite the presence of underlying fast and slow inhibitory postsynaptic potentials (IPSPs). DTNB decreased EPSPs to 48 +/- 12% (n = 5) of control. 5. Since a decrease of the NMDAR-mediated response by +/- 50% is sufficient to suppress the late part of the burst, we suggest that epileptiform activity can be controlled by manipulation of the redox sites of NMDAR. Our observations raise the possibility of developing new anticonvulsant drugs that would spare alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid-R (AMPAR)-mediated synaptic responses and decrease NMDAR-mediated synaptic transmission without blocking it completely.

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Decreased neuronal excitability in hippocampal neurons of mice exposed to cyclic hypoxia

Journal of Applied Physiology ◽

10.1152/jappl.2001.91.3.1245 ◽

2001 ◽

Vol 91 (3) ◽

pp. 1245-1250 ◽

Cited By ~ 37

Author(s):

Xiang Q. Gu ◽

Gabriel G. Haddad

Keyword(s):

Membrane Potential ◽

Hippocampal Neurons ◽

Neuronal Excitability ◽

Resting Membrane Potential ◽

Ca1 Neurons ◽

Recovery From Inactivation ◽

Channel Expression ◽

Steady State Inactivation ◽

And Function

To study the physiological effects of chronic intermittent hypoxia on neuronal excitability and function in mice, we exposed animals to cyclic hypoxia for 8 h daily (12 cycles/h) for ∼4 wk, starting at 2–3 days of age, and examined the properties of freshly dissociated hippocampal neurons in vitro. Compared with control (Con) hippocampal CA1 neurons, exposed (Cyc) neurons showed action potentials (AP) with a smaller amplitude and a longer duration and a more depolarized resting membrane potential. They also have a lower rate of spontaneous firing of AP and a higher rheobase. Furthermore, there was downregulation of the Na+ current density in Cyc compared with Con neurons (356.09 ± 54.03 pA/pF in Cyc neurons vs. 508.48 ± 67.30 pA/pF in Con, P < 0.04). Na+ channel characteristics, including activation, steady-state inactivation, and recovery from inactivation, were similar in both groups. The deactivation rate, however, was much larger in Cyc than in Con (at −100 mV, time constant for deactivation = 0.37 ± 0.04 ms in Cyc neurons and 0.18 ± 0.01 ms in Con neurons). We conclude that the decreased neuronal excitability in mice neurons treated with cyclic hypoxia is due, at least in part, to differences in passive properties (e.g., resting membrane potential) and in Na+ channel expression and/or regulation. We hypothesize that this decreased excitability is an adaptive response that attempts to decrease the energy expenditure that is used for adjusting disturbances in ionic homeostasis in low-O2conditions.

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Ventral Tegmental Area GABA, glutamate, and glutamate-GABA neurons are heterogenous in their electrophysiological and pharmacological properties

10.1101/2020.07.22.216093 ◽

2020 ◽

Author(s):

Jorge Miranda-Barrientos ◽

Ian Chambers ◽

Smriti Mongia ◽

Bing Liu ◽

Hui-Ling Wang ◽

...

Keyword(s):

Ventral Tegmental Area ◽

Viral Vectors ◽

Ex Vivo ◽

Glutamate Transporter ◽

Receptor Activation ◽

Firing Frequency ◽

Dopamine Neurons ◽

Gaba Neurons ◽

Double Transgenic Mouse ◽

Ventral Tegmental

AbstractThe ventral tegmental area (VTA) contains dopamine neurons intermixed with GABA-releasing (expressing vesicular GABA transporter, VGaT), glutamate-releasing (expressing vesicular glutamate transporter, VGluT2), and co-releasing (co-expressing VGaT and VGluT2) neurons. By delivering INTRSECT viral vectors into VTA of double vglut2-Cre/vgat-Flp transgenic mice, we targeted specific VTA cell populations for ex vivo recordings. We found that VGluT2+ VGaT− and VGluT2+ VGaT+ neurons on average had relatively hyperpolarized resting membrane voltage, greater rheobase, and lower spontaneous firing frequency compared to VGluT2− VGaT+ neurons, suggesting that VTA glutamate-releasing and glutamate-GABA co-releasing neurons require stronger excitatory drive to fire than GABA-releasing neurons. In addition, we detected expression of Oprm1mRNA (encoding μ opioid receptors, MOR) in VGluT2+ VGaT− and VGluT2− VGaT+ neurons, and their hyperpolarization by the MOR agonist DAMGO. Collectively, we demonstrate the utility of the double transgenic mouse to access VTA glutamate, glutamate-GABA and GABA neurons, and show some electrophysiological heterogeneity among them.Impact StatementSome physiological properties of VTA glutamate-releasing and glutamate-GABA co-releasing neurons are distinct from those of VTA GABA-releasing neurons. μ-opioid receptor activation hyperpolarizes some VTA glutamate-releasing and some GABA-releasing neurons.

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Differential impact of Kv8.2 loss on rod and cone signaling and degeneration.

10.1101/2021.07.05.451197 ◽

2021 ◽

Author(s):

Shivangi M Inamdar ◽

Colten K Lankford ◽

Deepak Poria ◽

Joseph G Laird ◽

Eduardo Solessio ◽

...

Keyword(s):

Membrane Potential ◽

Ex Vivo ◽

Light Stimulus ◽

Resting Membrane Potential ◽

Cone Dystrophy ◽

Extracellular Potassium ◽

Delayed Response ◽

Low Frequencies ◽

Ionic Imbalance

The voltage-gated potassium channel responsible for controlling photoreceptor signaling is a heteromeric complex of Kv2.1 subunits with a regulatory Kv8.2 subunit. Kv2.1/Kv8.2 channels are localized to the photoreceptor inner segment and carry IKx, largely responsible for setting the photoreceptor resting membrane potential. Mutations in Kv8.2 result in childhood-onset Cone Dystrophy with Supernormal Rod Response (CDSRR). We generated a Kv8.2 knockout (KO) mouse and examined retinal signaling and photoreceptor degeneration to gain deeper insight into the complex phenotypes of this disease. Using electroretinograms we show that there is a tradeoff between delayed or reduced signaling from rods depending on the intensity of the light stimulus, consistent with reduced capacity for light-evoked changes in membrane potential. The delayed response was not seen ex vivo where extracellular potassium levels are the same, so we conclude the in vivo alteration is influenced by ionic imbalance. We observed mild retinal degeneration. Signaling from cones was reduced but there was no loss of cone density. Loss of Kv8.2 altered responses to flickering light with responses attenuated at high frequencies and altered in shape at low frequencies. The Kv8.2 KO line on an all-cone retina background had reduced cone signaling associated with degeneration. We conclude that Kv8.2 is required by rods and cones for responding to dynamic changes in lighting. The timing and cell type affected by degeneration is different in the mouse and human but there is a window of time in both for therapeutic intervention.

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Abstract 338: Neuronal Responses to Hypoxic Stress in Mouse Cognitive Cortex

Circulation Research ◽

10.1161/res.111.suppl_1.a338 ◽

2012 ◽

Vol 111 (suppl_1) ◽

Author(s):

Guanghong Ding ◽

Ruqing Liu ◽

Dongman Chao ◽

Ying Xia

Keyword(s):

Young Adult ◽

Cortical Neurons ◽

Neuronal Excitability ◽

Membrane Conductance ◽

Pyramidal Cells ◽

Resting Membrane Potential ◽

Hypoxic Stress ◽

Open Probability ◽

Whole Cell ◽

Hypoxic Conditions

Cardiovascular disorders may cause hypoxic/ischemic injury to the cortex, leading to dysfunction of cortical cognition with a worse outcome in the older population. Although damage to cognitive cortex has been implicated in cognitive dysfunctions, the exact pathophysiology of a hypoxic/ischemic insult to this region is not well understood yet at the cellular and molecular levels. We used the whole-cell patch clamp in mouse slice preparations to characterize the basic electrophysiological properties of pyramidal cells of prefrontal cortex (PFC) and determine the response of these neurons to hypoxic stress in young adult (24±4 days old, n=7) and relatively older (42±12 days old, n=17) mice. In relatively older adult neurons (42±12 days old, n=17), the currents evoked by stepping from -140 mV to 20 mV (IV, holding at -60 mV) were shifted upward by hypoxic stress, indicating an increase in membrane conductance. Whole cell currents (holding at -60 mV) were found to decrease with a 3-min hypoxia, while increased with 5-min and 7-min hypoxia, suggesting that prolonged hypoxia increased open probability of ion channels. Resting membrane potential (MP) shifted upward in all three hypoxic conditions with a larger change seen with the longer duration of hypoxia. Action potentials (APs) showed the same tendency with 7-min hypoxia, showing a significant reduction in the number of hypoxia-stimulated pulses. However, all hypoxic conditions blocked the APs generation, suggesting hypoxic inhibition of neuronal excitability. In contrast, hypoxia-induced IV was apparently reversed in the younger neurons as compared to the older neurons, especially after 5 minutes of hypoxia (99.26 pA vs. -14.91 pA at -100 mV, P<0.01). Interestingly, even in normoxic condition, MP was significantly larger in young adult neurons (-74.38 mV) than in the older ones (-64.67 mV) (P<0.001). These data suggest that younger pyramidal cells of PFC are less vulnerable to hypoxic stress. Moreover, our preliminary data showed that activation of the delta-opioid receptors (DOR) inhibited the hypoxic response in the older neurons. Since acupuncture induces DOR up-regulation in the cortex, it is likely that acupuncture protects cortical neurons against hypoxic/ischemic stress via the DOR mechanism.

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TREK-1 K+ channels couple angiotensin II receptors to membrane depolarization and aldosterone secretion in bovine adrenal glomerulosa cells

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00223.2004 ◽

2004 ◽

Vol 287 (6) ◽

pp. E1154-E1165 ◽

Cited By ~ 37

Author(s):

Judith A. Enyeart ◽

Sanjay J. Danthi ◽

John J. Enyeart

Keyword(s):

Membrane Potential ◽

Angiotensin Ii ◽

Adrenal Cortex ◽

Receptor Activation ◽

Membrane Depolarization ◽

Resting Membrane Potential ◽

Zona Fasciculata ◽

Ang Ii ◽

Aldosterone Secretion ◽

Wide Range

Bovine adrenal glomerulosa (AZG) cells were shown to express bTREK-1 background K+ channels that set the resting membrane potential and couple angiotensin II (ANG II) receptor activation to membrane depolarization and aldosterone secretion. Northern blot and in situ hybridization studies demonstrated that bTREK-1 mRNA is uniformly distributed in the bovine adrenal cortex, including zona fasciculata and zona glomerulosa, but is absent from the medulla. TASK-3 mRNA, which codes for the predominant background K+ channel in rat AZG cells, is undetectable in the bovine adrenal cortex. In whole cell voltage clamp recordings, bovine AZG cells express a rapidly inactivating voltage-gated K+ current and a noninactivating background K+ current with properties that collectively identify it as bTREK-1. The outwardly rectifying K+ current was activated by intracellular acidification, ATP, and superfusion of bTREK-1 openers, including arachidonic acid (AA) and cinnamyl 1–3,4-dihydroxy-α-cyanocinnamate (CDC). Bovine chromaffin cells did not express this current. In voltage and current clamp recordings, ANG II (10 nM) selectively inhibited the noninactivating K+ current by 82.1 ± 6.1% and depolarized AZG cells by 31.6 ± 2.3 mV. CDC and AA overwhelmed ANG II-mediated inhibition of bTREK-1 and restored the resting membrane potential to its control value even in the continued presence of ANG II. Vasopressin (50 nM), which also physiologically stimulates aldosterone secretion, inhibited the background K+ current by 73.8 ± 9.4%. In contrast to its potent inhibition of bTREK-1, ANG II failed to alter the T-type Ca2+ current measured over a wide range of test potentials by using pipette solutions of identical nucleotide and Ca2+-buffering compositions. ANG II also failed to alter the voltage dependence of T channel activation under these same conditions. Overall, these results identify bTREK-1 K+ channels as a pivotal control point where ANG II receptor activation is transduced to depolarization-dependent Ca2+ entry and aldosterone secretion.

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Electrophysiological Alterations of Pyramidal Cells and Interneurons of the CA1 Region of the Hippocampus in a Novel Mouse Model of Dravet Syndrome

Genetics ◽

10.1534/genetics.120.303399 ◽

2020 ◽

Vol 215 (4) ◽

pp. 1055-1066

Author(s):

David A. Dyment ◽

Sarah C. Schock ◽

Kristen Deloughery ◽

Minh Hieu Tran ◽

Kerstin Ure ◽

...

Keyword(s):

Membrane Potential ◽

Mouse Model ◽

Hippocampal Slices ◽

Pyramidal Cells ◽

Epileptic Encephalopathy ◽

Dravet Syndrome ◽

Resting Membrane Potential ◽

Sodium Currents ◽

Spike Amplitude ◽

Induced Neurons

Dravet syndrome is a developmental epileptic encephalopathy caused by pathogenic variation in SCN1A. To characterize the pathogenic substitution (p.H939R) of a local individual with Dravet syndrome, fibroblast cells from the individual were reprogrammed to pluripotent stem cells and differentiated into neurons. Sodium currents of these neurons were compared with healthy control induced neurons. A novel Scn1aH939R/+ mouse model was generated with the p.H939R substitution. Immunohistochemistry and electrophysiological experiments were performed on hippocampal slices of Scn1aH939R/+ mice. We found that the sodium currents recorded in the proband-induced neurons were significantly smaller and slower compared to wild type (WT). The resting membrane potential and spike amplitude were significantly depolarized in the proband-induced neurons. Similar differences in resting membrane potential and spike amplitude were observed in the interneurons of the hippocampus of Scn1aH939R/+ mice. The Scn1aH939R/+ mice showed the characteristic features of a Dravet-like phenotype: increased mortality and both spontaneous and heat-induced seizures. Immunohistochemistry showed a reduction in amount of parvalbumin and vesicular acetylcholine transporter in the hippocampus of Scn1aH939R/+ compared to WT mice. Overall, these results underline hyper-excitability of the hippocampal CA1 circuit of this novel mouse model of Dravet syndrome which, under certain conditions, such as temperature, can trigger seizure activity. This hyper-excitability is due to the altered electrophysiological properties of pyramidal neurons and interneurons which are caused by the dysfunction of the sodium channel bearing the p.H939R substitution. This novel Dravet syndrome model also highlights the reduction in acetylcholine and the contribution of pyramidal cells, in addition to interneurons, to network hyper-excitability.

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Analgesic α-conotoxins modulate GIRK1/2 channels via GABAB receptor activation and reduce neuroexcitability

10.1101/2020.12.02.407627 ◽

2020 ◽

Author(s):

Anuja R. Bony ◽

Jeffrey R. McArthur ◽

Rocio K. Finol-Urdaneta ◽

David J. Adams

Keyword(s):

G Protein ◽

Neuronal Excitability ◽

Gabab Receptor ◽

Receptor Activation ◽

Resting Membrane Potential ◽

Membrane Hyperpolarization ◽

Analgesic Effects ◽

Inwardly Rectifying ◽

G Protein Coupled

AbstractActivation of G protein-coupled inwardly rectifying potassium (GIRK or Kir3) channels leads to membrane hyperpolarization and dampening of neuronal excitability. Here we show that the analgesic α-conotoxin Vc1.1 potentiates inwardly rectifying K+ currents (IKir) mediated through native and recombinant GIRK1/2 channels by activation of the G protein-coupled GABAB receptor (GABABR) via a Pertussis toxin (PTX)-sensitive G protein. Recombinant co-expression of human GIRK1/2 subunits and GABABR in HEK293T cells resulted in a Ba2+-sensitive IKir potentiated by baclofen and Vc1.1 which was inhibited by PTX, intracellular GDP-β-S, or the GABABR-selective antagonist CGP 55845. In adult mouse DRG neurons, GABABR-dependent GIRK channel potentiation by Vc1.1 and baclofen hyperpolarizes the cell resting membrane potential with concomitant reduction of excitability consistent with Vc1.1 and baclofen analgesic effects in vivo. This study provides new insight into Vc1.1 as an allosteric agonist for GABABR-mediated potentiation of GIRK channels and may aid in the development of novel non-opioid treatments for chronic pain.

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MicroRNA Biophysically Modulates Cardiac Action Potential by Direct Binding to Ion Channel

Circulation ◽

10.1161/circulationaha.120.050098 ◽

2021 ◽

Vol 143 (16) ◽

pp. 1597-1613 ◽

Cited By ~ 1

Author(s):

Dandan Yang ◽

Xiaoping Wan ◽

Adrienne T. Dennis ◽

Emre Bektik ◽

Zhihua Wang ◽

...

Keyword(s):

Membrane Potential ◽

Rna Interference ◽

Action Potential ◽

Ion Channel ◽

Cardiac Electrophysiology ◽

Ex Vivo ◽

Proximity Ligation Assay ◽

Resting Membrane Potential ◽

Seed Region ◽

Single Nucleotide

Background: MicroRNAs (miRs) play critical roles in regulation of numerous biological events, including cardiac electrophysiology and arrhythmia, through a canonical RNA interference mechanism. It remains unknown whether endogenous miRs modulate physiologic homeostasis of the heart through noncanonical mechanisms. Methods: We focused on the predominant miR of the heart (miR1) and investigated whether miR1 could physically bind with ion channels in cardiomyocytes by electrophoretic mobility shift assay, in situ proximity ligation assay, RNA pull down, and RNA immunoprecipitation assays. The functional modulations of cellular electrophysiology were evaluated by inside-out and whole-cell patch clamp. Mutagenesis of miR1 and the ion channel was used to understand the underlying mechanism. The effect on the heart ex vivo was demonstrated through investigating arrhythmia-associated human single nucleotide polymorphisms with miR1-deficient mice. Results: We found that endogenous miR1 could physically bind with cardiac membrane proteins, including an inward-rectifier potassium channel Kir2.1. The miR1–Kir2.1 physical interaction was observed in mouse, guinea pig, canine, and human cardiomyocytes. miR1 quickly and significantly suppressed I K1 at sub–pmol/L concentration, which is close to endogenous miR expression level. Acute presence of miR1 depolarized resting membrane potential and prolonged final repolarization of the action potential in cardiomyocytes. We identified 3 miR1-binding residues on the C-terminus of Kir2.1. Mechanistically, miR1 binds to the pore-facing G-loop of Kir2.1 through the core sequence AAGAAG, which is outside its RNA interference seed region. This biophysical modulation is involved in the dysregulation of gain-of-function Kir2.1–M301K mutation in short QT or atrial fibrillation. We found that an arrhythmia-associated human single nucleotide polymorphism of miR1 (hSNP14A/G) specifically disrupts the biophysical modulation while retaining the RNA interference function. It is remarkable that miR1 but not hSNP14A/G relieved the hyperpolarized resting membrane potential in miR1-deficient cardiomyocytes, improved the conduction velocity, and eliminated the high inducibility of arrhythmia in miR1-deficient hearts ex vivo. Conclusions: Our study reveals a novel evolutionarily conserved biophysical action of endogenous miRs in modulating cardiac electrophysiology. Our discovery of miRs’ biophysical modulation provides a more comprehensive understanding of ion channel dysregulation and may provide new insights into the pathogenesis of cardiac arrhythmias.

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