No abstract

The Ca 2+ -activated SK4 K + channel is gated by Ca 2+ –calmodulin (CaM) and is expressed in immune cells, brain, and heart. A cryoelectron microscopy (cryo-EM) structure of the human SK4 K + channel recently revealed four CaM molecules per channel tetramer, where the apo CaM C-lobe and the holo CaM N -lobe interact with the proximal carboxyl terminus and the linker S4–S5, respectively, to gate the channel. Here, we show that phosphatidylinositol 4-5 bisphosphate (PIP2) potently activates SK4 channels by docking to the boundary of the CaM-binding domain. An allosteric blocker, BA6b9, was designed to act to the CaM–PIP2-binding domain, a previously untargeted region of SK4 channels, at the interface of the proximal carboxyl terminus and the linker S4–S5. Site-directed mutagenesis, molecular docking, and patch-clamp electrophysiology indicate that BA6b9 inhibits SK4 channels by interacting with two specific residues, Arg191 and His192 in the linker S4–S5, not conserved in SK1–SK3 subunits, thereby conferring selectivity and preventing the Ca 2+ –CaM N -lobe from properly interacting with the channel linker region. Immunohistochemistry of the SK4 channel protein in rat hearts showed a widespread expression in the sarcolemma of atrial myocytes, with a sarcomeric striated Z-band pattern, and a weaker occurrence in the ventricle but a marked incidence at the intercalated discs. BA6b9 significantly prolonged atrial and atrioventricular effective refractory periods in rat isolated hearts and reduced atrial fibrillation induction ex vivo. Our work suggests that inhibition of SK4 K + channels by targeting drugs to the CaM–PIP2-binding domain provides a promising anti-arrhythmic therapy.

Robust electrical signal propagation in the form of action potentials (AP) is a hallmark of neuronal activity. While it is well established that changes to ionic gradients across the bilayer are responsible for AP propagation, the contributions of ionic diffusion and membrane morphological heterogeneity are not well understood. New experimental imaging methods have already suggested that spatial complexities of dendrites and the spatial dynamics of ionic species influence membrane voltage locally. However, it remains difficult to extract detailed voltage information at short time and length scales, which motivates a need for mathematical models that can incorporate these spatial complexities and predict and dissect voltage dynamics. Owing to the well-mixed assumption employed by popular models such as the Hodgkin-Huxley model, Morris-Lecar model, and cable theory the influence of morphology on voltage propagation cannot be studied. Therefore, building on these models, we construct a spatial model of AP propagation along dendrites which relaxes the well-mixed assumption. We explicitly model local ionic concentrations and their dynamics as influenced by reaction-diffusion and Hodgkin-Huxley type ion channel currents. The local transmembrane potential is determined by the local transmembrane ionic gradient via the Nernst potential. We compare membrane voltage dynamics from this new model to the traditional passive cable equation for dendrites of various sizes and configurations. Using our model, we find that a) membrane voltage propagation depends on the complex geometries of dendrites, b) the presence of internal organelles modulates membrane voltage propagation, and c) downstream signaling dynamics can affect AP propagation.