The presence of different currents in various tissues affects the complex role of BK channels in the regulation of excitability (Bailey et al. 2019). Therefore, defining the mutation's functional characterization seems mandatory. Numerous phenotypes were associated with KCNMA1 mutations. Particularly epilepsy, movement disorders, developmental delay, and cerebellar atrophy have been reported (Du et al. 2005; Zhang et al. 2015; Li et al. 2018; Tabarki et al. 2016). In the context of genetic heterogeneity, some of these variations show different BK current properties. BK channels play an important role in neuronal excitability via balancing potassium and calcium potential from depolarization to hyperpolarization. However, the exact mechanisms by which KCNMA1 variations effects BK channel activity remain unclear (Bailey et al. 2019). The understanding mechanistic explanation of the BK channel features and activities are not only crucial for neurological systems, but also muscular and osteoblastic activity (Jaggar et al. 2000; Tricarico et al. 2017; Hei et al. 2016).
As previously accounted, BK channel activity decreases the excitation feature of synaptic activity at neuromuscular junctions (Robitaille et al. 1993). Interestingly BK channels have a dual effect on neurons both for excitation and inhibition. Therefore, finding BK current activity shows different expression patterns for suppression to induction. For instance, Du et al. showed a missense variation, Asp434Gly in the regulator of conductance for K+ (RCK) domain in KCNMA1. In their study, Du et al. evaluated a large family with 13 individuals. Among them, one patient had only epilepsy, seven patients had paroxysmal dyskinesia and five patients had epilepsy with paroxysmal dyskinesia. Detected variations exhibited autosomal dominant inheritance patterns (Du et al. 2005). Latterly, the same mutation was detected and analyzed with different materials by other study groups (Diez-Sampedro et al. 2006; Wang et al. 2009; Yang et al. 2010). They demonstrate that the variation causes increased current activity, and they speculated a potential mechanism to account for disease in terms of this very specific variation. Briefly, this mechanism is G-V shift to hyperpolarized potentials, increased open probability, faster activation, slower deactivation, and increased Ca2+ sensitivity (Bailey et al. 2019). In contrast to these findings, Liang et al. reported three unrelated patients who had clinical heterogeneity, i.e. multiple malformation syndromes, such as facial abnormalities, global developmental delay, axial hypotonia, and visceral malformations. Also, one patient had a movement disorder, and two of the three had epileptic seizures (Liang et al. 2019). Similar to R458X reported by Yesil et.al, the variations C413Y, N449fs, and I663V reported by Liang et al. are placed in the RCK1 domain which is a binding site for Ca2+ (Liang et al. 2019). The clinical comparison of the patient carrying biallelic mutations C413Y/N449fs presented in and the patient presented by Yesil et. al. had developmental/intellectual deficiency and cerebellar ataxia in common however the patient reported by Yesil et. al had also dyskinesia and epilepsy phenotype (Yesil et al. 2018). Liang et al. observed that C413Y is a LoF variant that inhibits the function of the BK channel substantially and likely reduces the activation and macroscopic current amplitude of the BK channel significantly at calcium concentrations ranging from nominal 0 µM–10 Μm. Moreover, the frameshift variant p.(Asn449fs) did not elicit any potassium current under the voltage stimulus from − 160 mV–160 mV at a 10 µM calcium concentration suggesting that it is also a LoF variant.
In this study report, we conducted patch-clamp recordings on WT and R458X mutant cells. To understand how a truncating mutation abolishes function and the electrophysiological outcome, a patch-clamp with cell-attached recordings is a direct method. We measured current density from the ratio of peak current amplitude to the cell membrane capacitance (picoamperes per picofarad (pA/pF)) under a stable pH. We found out that under a stable pH the mutant type showed an increasing sequence of actions when compared to the wild type. While the stimulus continued, changes in the potassium channel activation might have altered downstream potential as well. It seems that the increased current and thus the hyperpolarized state -of the channel firstly alters the function but negatively affects the viability of the cell later on (Fig. 1, 2). The dual phenotype with epilepsy/dyskinesia and cerebellar atrophy, may be explained with this mechanism in such severe gain of function mutations. Although gain-of-function have been generally shown in missense mutations nonsense mutations with a gain of function consequence have also been reported in the literature (Ho et al. 2007; Kuehnet al. 2017 ).
Taken together, the nature of the BK-channel and its effect on organisms, show a giant heterogeneity, from transcript variation to cellular electrical activity. Both the literature and this study have shown that the, changes in the RCK domain of BK channels directly affect protein function (Du et al. 2005; Zhang et al. 2010; Kim et al. 2008). This is the first functional study observing an increased current in the KCNMA1 gene resulting from a truncating mutation. In conclusion, due to its complexity of their nature every causative variant in the channel genes should be evaluated in terms of functional characterization, particularly electrophysiological analyzes.