Holstein-Rathlou, N.-H. | Search bioRxiv

bioRxiv2024

Ephaptic coupling enables action potential conduction

BackgroundCardiac action potentials are believed to propagate solely by electrotonic transmission via gap junctions. In the alternative theory of ephaptic transmission, changes in extracellular electrical fields and ion concentrations may enable a cardiomyocyte to activate its neighbors. The involvement of ephaptic mechanisms remain highly disputed and has been discounted due to the reported inability of an isolated cardiomyocyte to activate another. MethodsIsolated rat cardiomyocytes were subjected to whole cell patch clamp. Transmission of action potentials was tested in current clamp in isolated cardiomyocytes pushed together and in native cardiomyocyte pairs. Sodium channel activation and inactivation was tested in voltage clamp. ResultsIsolated cardiomyocytes were pushed together end-to-end and one stimulated to fire an action potential. In 10 attempts, 2 pairs showed successful transmission of action potentials, showing that ephaptic transmission is possible. In native pairs, subthreshold depolarization was transferred between cardiomyocytes of native electrically coupled pairs, however, only after a delay, which is not be explained from electrotonic theory. Once action potentials were elicited in one cardiomyocyte of a native pair, the neighbor was activated with delays under 2 msec. The activation delay did not correlate with gap junctional coupling, again suggesting the presence of an alternative mechanism. In voltage clamp of native cardiomyocyte pairs, one cardiomyocyte was held at -90 mV while depolarizing pulses were applied to its neighbor. Consistently, activation of sodium channels in the depolarized cell led to activation of sodium channels in the hyperpolarized cell, which fits well with ephaptic but not electrotonic theory. Ephaptic transmission requires a shielded local domain. To separate this, we measured steady state inactivation in native pairs. As expected, all channels inactivated at -40 mV. However, when maintaining a potential of - 40 mV for an extended period, sodium current elicited at 0 mV re-emerged by de-inactivation. The de-inactivation at -40 mV support that a shielded domain exists where potassium currents can hyperpolarize the local potential. ConclusionsWe show for the first time that ephaptic transmission is possible, that it is most likely in operation, also in the presence of gap junctional coupling, and finally our study suggests that data obtained in single cells may fail to reproduce key features of the electrophysiology of cardiomyocytes that interact with neighboring cardiomyocytes.

physiology↗

bioRxiv2021

Blood flow synchronization in renal microcirculation - a high-resolution imaging study.

AimsInternephron signalling and interaction are fundamental for kidney function. Earlier studies have shown that nephrons signal to each other over short distances and adjust their activity accordingly. Micropuncture experiments revealed synchronous clusters of 2-3 nephrons formed from such interactions, while imaging and modelling results suggested the possibility of larger clusters. Such clusters are expected to play an important role in renal autoregulation, but their presence has not been confirmed and their size has not been estimated. In this study, we present methodology for high resolution renal blood flow imaging and apply it to estimate frequency and phase angle differences in kidney blood vessels under normal conditions and after administration of the vasoactive agents angiotensin II and acetylcholine. Methods and resultsTo resolve signals from separate arterioles in a sufficiently large field of view, we developed a method for renal laser speckle contrast imaging. Our setup provides imaging of blood flow in the kidney cortex with a limit of image resolution at 0.8m per pixel and imaging frequency of 160Hz. We used the method to record from 1.5x1.5 mm2 sections of the renal surface in anaesthetised Sprague-Dawley rats in unstimulated conditions and during IV infusion of the vasoconstrictor angiotensin II or the vasodilator acetylcholine. In each section, we resolved and segmented 94.8{+/-}15.66 individual arterioles and venules, and analyzed blood flow using wavelet spectral analysis to identify clusters of synchronized blood vessels. ConclusionsWe observed spatial and temporal evolution of blood vessel clusters of various sizes, including the formation of large (>90 vessels) long-lived clusters (>10 periods) locked at the frequency of the tubular glomerular feedback (TGF) mechanism. The analysis showed that synchronization patterns and thus the co-operative dynamics of nephrons change significantly when either of the vasoactive agents is administered. On average, synchronization was stronger (larger clusters, longer duration) with angiotensin II administration than in the unstimulated state or with acetyl choline. While it weakens with distance, increased synchronization duration spanned the whole field of view, and likely, beyond it. Neighbouring vessels tend to demonstrate in-phase synchronization, especially in the vasoconstricted condition, which is expected to cause locally increased pressure variation. Our results confirm both the presence of the local synchronization in the renal microcirculatory blood flow and the fact that it changes depending on the condition of the vascular network and the blood pressure, which might have further implications for the role of such synchronization in pathologies development.

physiology↗