BK Channels Are Activated by Functional Coupling With L-Type Ca2+ Channels in Cricket Myocytes

Large-conductance calcium (Ca2+)-activated potassium (K+) (BK) channel activation is important for feedback control of Ca2+ influx and cell excitability during spontaneous muscle contraction. To characterize endogenously expressed BK channels and evaluate the functional relevance of Ca2+ sources leading to BK activity, patch-clamp electrophysiology was performed on cricket oviduct myocytes to obtain single-channel recordings. The single-channel conductance of BK channels was 120 pS, with increased activity resulting from membrane depolarization or increased intracellular Ca2+ concentration. Extracellular application of tetraethylammonium (TEA) and iberiotoxin (IbTX) suppressed single-channel current amplitude. These results indicate that BK channels are endogenously expressed in cricket oviduct myocytes. Ca2+ release from internal Ca2+ stores and Ca2+ influx via the plasma membrane, which affect BK activity, were investigated. Extracellular Ca2+ removal nullified BK activity. Administration of ryanodine and caffeine reduced BK activity. Administration of L-type Ca2+ channel activity regulators (Bay K 8644 and nifedipine) increased and decreased BK activity, respectively. Finally, the proximity between the L-type Ca2+ channel and BK was investigated. Administration of Bay K 8644 to the microscopic area within the pipette increased BK activity. However, this increase was not observed at a sustained depolarizing potential. These results show that BK channels are endogenously expressed in cricket oviduct myocytes and that BK activity is regulated by L-type Ca2+ channel activity and Ca2+ release from Ca2+ stores. Together, these results show that functional coupling between L-type Ca2+ and BK channels may underlie the molecular basis of spontaneous rhythmic contraction.

Synthesis and Biological Activity of a Bis-steroid-methanocyclobutanaphthalene- dione Derivative against Ischemia/Reperfusion Injury via Calcium Channel Activation

Background: There is some experimental data on the effect exerted by some steroid derivatives against ischemia/reperfusion injury; however, the molecular mechanism is very confusing, perhaps this phenomenon could be due to the protocols used and/or differences in the chemical structure of each one of the steroid derivatives. Objective: The aim of this study was to synthesize a new bis-steroid-methanocyclobutanaphthalene- dione derivative using some tools chemical. Methodology: The biological activity exerted by the bis-steroid-methanocyclobutanaphthalene- dione derivative against ischemia/reperfusion injury was evaluated in an isolated heart model using noradrenaline, milrinone, dobutamine, levosimendan, and Bay-K- 8644 as controls. In addition, other alternative experiments were carried out to evaluate the biological activity induced by the bis-steroid-methanocyclobuta-naphthalene-dione derivative against left ventricular pressure in the absence or presence of nifedipine. Results: The results showed that 1) the bis-steroid-methanocyclobuta-naphthalene-dione derivative significantly decreases the ischemia-reperfusion injury translated as a decrease in the the infarct area in a similar manner to levosimendan drug; 2) both bis-steroidmethanocyclobuta- naphthalene-dione and Bay-K-8644 increase the left ventricular pressure and 3) the biological activity exerted by bis-steroid-methanocyclobuta-naphthalenedione derivative against left ventricular pressure is inhibited by nifedipine. Conclusion: In conclusion, the bis-steroid-methanocyclobuta-naphthalene-dione derivative decreases the area of infarction and increases left ventricle pressure via calcium channels activation; this phenomenon could constitute a new therapy for ischemia/reperfusion injury.

L-type calcium channel contributions to intrinsic excitability and synaptic activity during basolateral amygdala postnatal development

The amygdala contributes toward emotional processes such as fear, anxiety, and social cognition. Furthermore, evidence suggests that increased excitability of basolateral amygdala (BLA) principal neurons underlie certain neuropsychiatric disorders. Gain-of-function mutations in neuronal L-type calcium channels (LTCCs) are linked to neurodevelopmental diseases, including autism spectrum disorders (ASDs). While LTCCs are expressed throughout the BLA, direct evidence for increased LTCC activity affecting BLA excitability and potentially contributing to disease pathophysiology is lacking. In this study, we utilized a pharmacological approach to examine the contributions of LTCCs to BLA principal cell excitability and synaptic activity at immature (postnatal day 7, P7) and juvenile (P21) developmental stages. Acute upregulation of LTCC activity in brain slices by application of the agonist ( S)-Bay K 8644 resulted in increased intrinsic excitability properties including firing frequency response, plateau potential, and spike-frequency adaptation selectively in P7 neurons. Contrastingly, for P21 neurons, the main effect of ( S)-Bay K 8644 was to enhance burst firing. ( S)-Bay K 8644 increased spontaneous inhibitory synaptic currents at both P7 and P21 but did not affect evoked synaptic currents at either stage. ( S)-Bay K 8644 did not alter P7 spontaneous excitatory synaptic currents, although it increased current amplitude in P21 neurons. Overall, the results provide support for the notion that alteration of LTCC activity at specific periods of early brain development may lead to functional alterations to neuronal network activity and subsequently contribute to underlying mechanisms of amygdala-related neurological disorders. NEW & NOTEWORTHY The role of L-type calcium channels (LTCCs) in regulating neuronal electrophysiological properties during development remains unclear. We show that in basolateral amygdala principal neurons, an increase of LTCC activity alters both neuronal excitability and synaptic activity. The results also provide evidence for the distinct contributions of LTCCs at different stages of neurodevelopment and shed insight into our understanding of LTCC dysfunction in amygdala-related neurological disorders.

Abstract 14727: Late Sodium Current Plays a Role in Ventricular Arrhythmias Associated With Increased Intracellular Calcium Concentration

Objective: An increase in intracellular calcium concentration is associated with the prolongation of action potential duration (APD) and polymorphic ventricular tachycardia (PVT). Recent studies indicated that late sodium current inhibitor is effective in preventing ventricular arrhythmias in patients with long QT syndrome 8. The objective of the study was to determine the role of late sodium current in calcium related ventricular arrhythmias. Methods: Hearts from New Zealand female rabbits were perfused in a Langendorff mode. The atrioventricular nodal area was thermally ablated to produce complete atrioventricular block, and then heart was paced at stated frequency. Multiple channel monophasic action potentials (MAP) and pseudo 12-lead electrocardiograms (ECGs) were recorded. Calcium transient and myocyte contraction were determined in rabbit ventricular myocytes. Results: Bay-K 8644 (10-300 nM) increased both epi- and endo- cardial MAPD90 of left ventricle in concentration dependent manners, from (176±6) to (222±13) ms, and (201±6) to (246±10) ms (n=15, p<0.05 vs control), respectively. In the presence of 1 nM ATX-II, Bay-K 8644 caused a greater prolongation of epi-MAPD90 which was increased from (182±6) to (342±21) ms (n=9, p<0.05 vs control). The prolongation of MAPD90 caused by Bay-K 8644 were reversed by 1 μM TTX in both absence and presence of ATX-II. Additionally, the incidence of PVT evoked by Bay-K 8644 was also greater in the presence compared to the absence of ATX-II. 200 nM Bay-K 8644 caused few arrhythmias in absence of ATX-II. In contrast, PVT occurred in 7/9 (77.78%) of hearts treated with 200 nM Bay-K 8644 in the presence of ATX-II. These arrhythmias could be abolished by 1 μM TTX in the continued presence of Bay-K 8644. TTX (1 μM) attenuated the increase by 200 nM Bay-K 8644 of intracellular calcium transient and myocyte contraction amplitude by 10.8% and 14.6%, respectively (n=6, p<0.05). Conclusion: Both endogenous and enhanced late sodium current contributes to the ventricular arrhythmias with increased intracellular calcium concentration. Inhibition of late sodium current may be effective in preventing or treating calcium overload-related ventricular arrhythmias and dysfunction of myocardial contraction.