Heart Rate and Extracellular Sodium and Potassium Modulation of Gap Junction Mediated Conduction in Guinea Pigs

Abstract

Background: Recent studies suggested that cardiac conduction in murine hearts with narrow perinexi and 50% reduced connexin43 (Cx43) expression is more sensitive to relatively physiological changes of extracellular potassium ([K+]o) and sodium ([Na+]o).Purpose: Determine whether similar [K+]o and [Na+]o changes alter conduction velocity (CV) sensitivity to pharmacologic gap junction (GJ) uncoupling in guinea pigs.Methods: [K+]o and [Na+]o were varied in Langendorff perfused guinea pig ventricles (Solution A: [K+]o = 4.56 and [Na+]o = 153.3 mM. Solution B: [K+]o = 6.95 and [Na+]o = 145.5 mM). Gap junctions were inhibited with carbenoxolone (CBX) (15 and 30 μM). Epicardial CV was quantified by optical mapping. Perinexal width was measured with transmission electron microscopy. Total and phosphorylated Cx43 were evaluated by western blotting.Results: Solution composition did not alter CV under control conditions or with 15μM CBX. Decreasing the basic cycle length (BCL) of pacing from 300 to 160 ms decreased CV uniformly with both solutions. At 30 μM CBX, a change in solution did not alter CV either longitudinally or transversely at BCL = 300 ms. However, reducing BCL to 160 ms caused CV to decrease more in hearts perfused with Solution B than A. Solution composition did not alter perinexal width, nor did it change total or phosphorylated serine 368 Cx43 expression. These data suggest that the solution dependent CV changes were independent of altered perinexal width or GJ coupling. Action potential duration was always shorter in hearts perfused with Solution B than A, independent of pacing rate and/or CBX concentration.Conclusions: Increased heart rate and GJ uncoupling can unmask small CV differences caused by changing [K+]o and [Na+]o. These data suggest that modulating extracellular ionic composition may be a novel anti-arrhythmic target in diseases with abnormal GJ coupling, particularly when heart rate cannot be controlled.

Revealing the Concealed Nature of Long-QT Type 3 Syndrome

“…Further, consistent with our prior work, 20, 25, 26 TEM micrographs illustrate the spatially restricted nature of intercellular clefts (Figure 2A). The high density of Na + channels localized at the ID suggests that extracellular cleft Na + depletion may be an important aspect of Na + -dependent signaling and demonstrates the need for computational modeling to account for the time-dependent dynamics of Na + influx, efflux, and diffusion at the intercellular cleft, especially in pathological settings of Na + channel mutations.…”

“…We first tested the hypothesis that ID expansion can prolong APD and unmask EADs in a drug-induced, guinea pig model of LQT3. Representative TEM micrographs and summary data from a blinded observer demonstrated that intercellular separation within the perinexus (Figure 2A) with ATXII was 18.3±0.7nm (95%-confidence interval (CI):17.0–19.6nm, blue), similar to previously published measurements in murine and guinea pig ventricular myocardium estimated by the same observer with similar perfusate compositions 20, 25, 26 . Mannitol significantly expanded w P to 24.4±0.7 nm (95%-CI:23.1–25.7nm, AIE, green; p=0.003).…”

“…The model predicts the Y1795C mutation slightly increased CV relative to WT at clefts widths greater than 20nm and less than 10nm. However, in the nominal cleft width range found in guinea pig and mouse ventricular tissue (10 – 20nm) 8, 26 , CV is predicted to be relatively similar for WT and mutant strands.…”

“…EpC in cardiac tissue has been investigated in prior computational studies 10, 16, 17, 19, 32 and recently by us experimentally 8, 20, 24–26, 42 . These studies have typically focused on how EpC influences conduction and is modulated by gap junctional coupling, extracellular ionic concentrations, and other factors.…”

“…We recognize that edema is a complex clinical pathology involving intra and extracellular volume expansion. Transmission electron microscopy (TEM) was performed as previously described 20, 25, 26 on 3 hearts per treatment. Additional details can be found in Supplementary Methods.…”