Nicholas C. Foeger

Homeostatic regulation of electrical excitability in physiological cardiac hypertrophy

Pathological biomechanical stresses cause cardiac hypertrophy, which is associated with QT prolongation and arrhythmias. Previous studies have demonstrated that repolarizing K+ current densities are decreased in pressure overload‐induced left ventricular hypertrophy, resulting in action potential and QT prolongation. Cardiac hypertrophy also occurs with exercise training, but this physiological hypertrophy is not associated with electrical abnormalities or increased arrhythmia risk, suggesting that repolarizing K+ currents are upregulated, in parallel with the increase in myocyte size, to maintain normal cardiac function. To explore this hypothesis directly, electrophysiological recordings were obtained from ventricular myocytes isolated from two mouse models of physiological hypertrophy, one produced by swim‐training of wild‐type mice and the other by cardiac‐specific expression of constitutively active phosphoinositide‐3‐kinase‐p110α (caPI3Kα). Whole‐cell voltage‐clamp recordings revealed that repolarizing K+ current amplitudes were higher in ventricular myocytes isolated from swim‐trained and caPI3Kα, compared with wild‐type, animals. The increases in K+ current amplitudes paralleled the observed cellular hypertrophy, resulting in normalized or increased K+ current densities. Electrocardiographic parameters, including QT intervals, as well as ventricular action potential waveforms in swim‐trained animals/myocytes were indistinguishable from controls, demonstrating preserved electrical function. Additional experiments revealed that inward Ca2+ current amplitudes/densities were also increased in caPI3Kα, compared with WT, left ventricular myocytes. The expression of transcripts encoding K+, Ca2+ and other ion channel subunits was increased in swim‐trained and caPI3Kα ventricles, in parallel with the increase in myocyte size and with the global increases in total cellular RNA expression. In contrast to pathological hypertrophy, therefore, the functional expression of repolarizing K+ (and depolarizing Ca2+) channels is increased with physiological hypertrophy, reflecting upregulation of the underlying ion channel subunit transcripts and resulting in increased current amplitudes and the normalization of current densities and action potential waveforms. Taken together, these results suggest that activation of PI3Kα signalling preserves normal myocardial electrical functioning and could be protective against the increased risk of arrhythmias and sudden death that are prevalent in pathological cardiac hypertrophy.

Interdependent Roles for Accessory KChIP2, KChIP3, and KChIP4 Subunits in the Generation of Kv4-Encoded IA Channels in Cortical Pyramidal Neurons

The rapidly activating and inactivating voltage-dependent outward K+ (Kv) current, IA, is widely expressed in central and peripheral neurons. IA has long been recognized to play important roles in determining neuronal firing properties and regulating neuronal excitability. Previous work demonstrated that Kv4.2 and Kv4.3 α-subunits are the primary determinants of IA in mouse cortical pyramidal neurons. Accumulating evidence indicates that native neuronal Kv4 channels function in macromolecular protein complexes that contain accessory subunits and other regulatory molecules. The K+ channel interacting proteins (KChIPs) are among the identified Kv4 channel accessory subunits and are thought to be important for the formation and functioning of neuronal Kv4 channel complexes. Molecular genetic, biochemical, and electrophysiological approaches were exploited in the experiments described here to examine directly the roles of KChIPs in the generation of functional Kv4-encoded IA channels. These combined experiments revealed that KChIP2, KChIP3, and KChIP4 are robustly expressed in adult mouse posterior (visual) cortex and that all three proteins coimmunoprecipitate with Kv4.2. In addition, in cortical pyramidal neurons from mice lacking KChIP3 (KChIP3−/−), mean IA densities were reduced modestly, whereas in mean IA densities in KChIP2−/− and WT neurons were not significantly different. Interestingly, in both KChIP3−/− and KChIP2−/− cortices, the expression levels of the other KChIPs (KChIP2 and 4 or KChIP3 and 4, respectively) were increased. In neurons expressing constructs to mediate simultaneous RNA interference-induced reductions in the expression of KChIP2, 3, and 4, IA densities were markedly reduced and Kv current remodeling was evident.

Stabilization of Kv4 protein by the accessory K+ channel interacting protein 2 (KChIP2) subunit is required for the generation of native myocardial fast transient outward K+ currents

•  The cytosolic K+ channel accessory subunit, K+ channel interacting protein 2 (KChIP2), was previously suggested to be critical in the generation of cardiac fast transient outward current (Ito,f) channels. •  The experiments presented here revealed the novel finding that targeted deletion of KChIP2 results in the complete loss of the Kv4.2 protein, although Kcnd2 (Kv4.2) transcript expression is not decreased in KChIP2−/− ventricles. •  In contrast, the slow transient outward current, Ito,s, is increased in KChIP2−/− left ventricular apex myocytes and ventricular action potential waveforms in KChIP2−/− and WT mice are not significantly different. •  These results demonstrate the critical role of KChIP2 in the stabilization of native Kv4 proteins and that the loss of the Kv4.2 protein underlies the elimination of Ito,f in KChIP2−/− myocytes. •  Taken together, the results here demonstrate that electrical remodelling compensates for the elimination of Ito,f, maintaining physiological action potential repolarization in mouse myocardium.

Co-assembly of Kv4 α Subunits with K+ Channel-interacting Protein 2 Stabilizes Protein Expression and Promotes Surface Retention of Channel Complexes

Members of the K+ channel-interacting protein (KChIP) family bind the distal N termini of members of the Shal subfamily of voltage-gated K+ channel (Kv4) pore-forming (α) subunits to generate rapidly activating, rapidly inactivating neuronal A-type (IA) and cardiac transient outward (Ito) currents. In heterologous cells, KChIP co-expression increases cell surface expression of Kv4 α subunits and Kv4 current densities, findings interpreted to suggest that Kv4·KChIP complex formation enhances forward trafficking of channels (from the endoplasmic reticulum or the Golgi complex) to the surface membrane. The results of experiments here, however, demonstrate that KChIP2 increases cell surface Kv4.2 protein expression (∼40-fold) by an order of magnitude more than the increase in total protein (∼2-fold) or in current densities (∼3-fold), suggesting that mechanisms at the cell surface regulate the functional expression of Kv4.2 channels. Additional experiments demonstrated that KChIP2 decreases the turnover rate of cell surface Kv4.2 protein by inhibiting endocytosis and/or promoting recycling. Unexpectedly, the experiments here also revealed that Kv4.2·KChIP2 complex formation stabilizes not only (total and cell surface) Kv4.2 but also KChIP2 protein expression. This reciprocal protein stabilization and Kv4·KChIP2 complex formation are lost with deletion of the distal (10 amino acids) Kv4.2 N terminus. Taken together, these observations demonstrate that KChIP2 differentially regulates total and cell surface Kv4.2 protein expression and Kv4 current densities.

Augmentation of Kv4.2-encoded currents by accessory dipeptidyl peptidase 6 and 10 subunits reflects selective cell surface Kv4.2 protein stabilization.

Background: Somatodendritic Kv4-encoded A-Type K+ current densities are enhanced by both cytosolic K+ channel interacting proteins (KChIPs) and transmembrane dipeptidyl peptidases (DPPs). Results: DPPs selectively stabilize cell surface Kv4 protein expression, whereas KChIPs stabilize total and surface Kv4 protein expression. Conclusion: DPPs regulate Kv4-encoded current densities through mechanisms distinct from the KChIPs. Significance: Multiple mechanisms determine functional Kv4 channel densities. Rapidly activating and inactivating somatodendritic voltage-gated K+ (Kv) currents, IA, play critical roles in the regulation of neuronal excitability. Considerable evidence suggests that native neuronal IA channels function in macromolecular protein complexes comprising pore-forming (α) subunits of the Kv4 subfamily together with cytosolic, K+ channel interacting proteins (KChIPs) and transmembrane, dipeptidyl peptidase 6 and 10 (DPP6/10) accessory subunits, as well as other accessory and regulatory proteins. Several recent studies have demonstrated a critical role for the KChIP subunits in the generation of native Kv4.2-encoded channels and that Kv4.2-KChIP complex formation results in mutual (Kv4.2-KChIP) protein stabilization. The results of the experiments here, however, demonstrate that expression of DPP6 in the mouse cortex is unaffected by the targeted deletion of Kv4.2 and/or Kv4.3. Further experiments revealed that heterologously expressed DPP6 and DPP10 localize to the cell surface in the absence of Kv4.2, and that co-expression with Kv4.2 does not affect total or cell surface DPP6 or DPP10 protein levels. In the presence of DPP6 or DPP10, however, cell surface Kv4.2 protein expression is selectively increased. Further addition of KChIP3 in the presence of DPP10 markedly increases total and cell surface Kv4.2 protein levels, compared with cells expressing only Kv4.2 and DPP10. Taken together, the results presented here demonstrate that the expression and localization of the DPP accessory subunits are independent of Kv4 α subunits and further that the DPP6/10 and KChIP accessory subunits independently stabilize the surface expression of Kv4.2.

Neuronal Voltage-Gated K+ (Kv) Channels Function in Macromolecular Complexes

Considerable evidence indicates that native neuronal voltage-gated K+ (Kv) currents reflect the functioning of macromolecular Kv channel complexes, composed of pore-forming (α)-subunits, cytosolic and transmembrane accessory subunits, together with regulatory and scaffolding proteins. The individual components of these macromolecular complexes appear to influence the stability, the trafficking, the localization and/or the biophysical properties of the channels. Recent studies suggest that Kv channel accessory subunits subserve multiple roles in the generation of native neuronal Kv channels. Additional recent findings suggest that Kv channel accessory subunits can respond to changes in intracellular Ca(2+) or metabolism and thereby integrate signaling pathways to regulate Kv channel expression and properties. Although studies in heterologous cells have provided important insights into the effects of accessory subunits on Kv channel expression/properties, it has become increasingly clear that experiments in neurons are required to define the physiological roles of Kv channel accessory and associated proteins. A number of technological and experimental hurdles remain that must be overcome in the design, execution and interpretation of experiments aimed at detailing the functional roles of accessory subunits and associated proteins in the generation of native neuronal Kv channels. With the increasing association of altered Kv channel functioning with neurological disorders, the potential impact of these efforts is clear.

Long-Term Outcomes Following a Single Corticosteroid Injection for Trigger Finger

BACKGROUND The outcomes of corticosteroid injection for trigger finger are well documented only with short-term follow-up. The purpose of this investigation was to determine the long-term effectiveness of a single injection and to examine predictors of success up to ten years after injection. METHODS This case series analyzed 366 first-time corticosteroid injections in flexor tendon sheaths from January 2000 to December 2007 with a minimum follow-up duration of five years. Two hundred and forty patients (66%) were female, 161 patients (44%) had multiple trigger fingers, and eighty-eight patients (24%) had diabetes at the time of injection. The primary outcome of treatment failure was defined as subsequent injection or surgical trigger finger release of the affected digit. Medical records were reviewed, and any patients without documented failure or a return office visit in 2012 to 2013 were contacted by telephone regarding symptom recurrence and the need for additional treatment. Kaplan-Meier analyses with log-rank test and Cox regression analysis assessed the effect of baseline patient and disease characteristics on injection success. RESULTS Forty-five percent of patients demonstrated long-term treatment success after a single injection. In the final regression model, the interaction of sex and the number of trigger fingers was the single predictor of treatment success. Exploring this association revealed a ten-year success rate of 56% for female patients presenting for the first time with a trigger finger compared with 35% in male patients presenting for the first time with a trigger finger, 39% in female patients with multiple trigger fingers, and 37% in male patients with multiple trigger fingers. Eighty-four percent of treatment failures occurred within the first two years following injection. Patient age, symptom type, and undifferentiated diabetes status were not predictive of treatment success. CONCLUSIONS Female patients presenting with their first trigger finger have the highest rate of long-term treatment success after a single corticosteroid injection. Patients who continue to experience symptom relief two years after injection are likely to maintain long-term success.

K+ Channel Interacting Proteins 2, 3 and 4 are Critical Components of Kv4 Channel Complexes in Cortical Pyramidal Neurons

The rapidly activating and inactivating voltage-gated K+ (Kv) current, IA, is critical for many neuronal functions, including repetitive firing and synaptic integration. Previous studies revealed that in cortical pyramidal neurons the majority of IA is encoded by Kv4.2 and Kv4.3 α-subunits. Little, however, is known about the functional roles of K+ Channel Interacting Proteins (KChIP) 1, 2, 3, and 4 in the generation of IA. Biochemical experiments revealed that KChIPs 2, 3 and 4 (2-4) co-immunoprecipitate with Kv4.2 in samples from mouse cortex suggesting roles for these three KChIPs in the generation of functional Kv4-encoded channels in cortical pyramidal neurons. Electrophysiological experiments conducted on cortical pyramidal neurons from mice (KChIP3-/-) harboring a targeted disruption of the KChIP3 locus revealed that IA densities and properties were similar to wild type neurons. Interestingly, in cortical samples from KChIP3-/- mice the protein levels of KChIP 2 and 4 were increased suggesting functional compensation for the loss of KChIP3. Similarly, in KChIP2-/- cortices KChIP3 and 4 protein levels were increased relative to wild type. Concurrently knocking down the expression of KChIPs 2-4 using RNAi constructs targeting each of the three KChIPs induced a reduction in IA density consistent with roles for KChIPs 2-4 in the generation of native Kv4-encoded IA channels. In cortical samples from Kv4.2-/- and Kv4.3-/- mice, the protein expression levels of KChIPs 2-4 were decreased. Additionally, in samples from mice lacking both Kv4.2 and Kv4.3 KChIP2-4 proteins were barely detectable. Taken together these results demonstrate that KChIPs 2-4 associate with Kv4.2 and Kv4.3 in cortical neurons, this association stabilizes KChIP proteins and, in addition, that KChIPs 2-4 are critical components of native Kv4 channels in cortical pyramidal neurons.