The transverse-axial tubular system (t-tubules) plays an essential role in excitation-contraction coupling in cardiomyocytes. Its remodelling is associated with various cardiac diseases. Numerous attempts were made to analyse characteristics essential for proper understanding of the t-tubules and their impact on cardiac cell function in health and disease. The currently available methodical approaches related to the fraction of the t-tubular membrane area produce diverse data. The widely used detubulation techniques cause irreversible cell impairment, thus, distinct cell samples have to be used for estimation of t-tubular parameters in untreated and detubulated cells. Our proposed alternative method is reversible and allows repetitive estimation of the fraction of t-tubular membrane (f t) in cardiomyocytes using short-term perfusion of the measured cell with a low-conductive isotonic sucrose solution. It results in a substantial increase in the electrical resistance of t-tubular lumen, thus, electrically separating the surface and t-tubular membranes. Using the whole-cell patch-clamp measurement and the new approach in enzymatically isolated rat atrial and ventricular myocytes, a set of data was measured and evaluated. The analysis of the electrical equivalent circuit resulted in the establishment of criteria for excluding measurements in which perfusion with a low conductivity solution did not affect the entire cell surface. As expected, the final average f t in ventricular myocytes (0.337 ± 0.017) was significantly higher than that in atrial myocytes (0.144 ± 0.015). The parameter f t could be estimated repetitively in a particular cell (0.345 ± 0.021 and 0.347 ± 0.023 in ventricular myocytes during the first and second sucrose perfusion, respectively). The new method is fast, simple, and leaves the measured cell intact. It can be applied in the course of experiments for which it is useful to estimate both the surface and t-tubular capacitance/area in a particular cell.

• Klíčová slova
• detubulation, membrane capacitance, new method, rat cardiomyocytes, sucrose, t-tubules,
• Publikační typ
• časopisecké články MeSH

Nicotine abuse is associated with variety of diseases including arrhythmias, most often atrial fibrillation (AF). Altered inward rectifier potassium currents including acetylcholine-sensitive current I K(Ach) are known to be related to AF pathogenesis. Since relevant data are missing, we aimed to investigate I K(Ach) changes at clinically relevant concentrations of nicotine. Experiments were performed by the whole cell patch clamp technique at 23 ± 1 °C on isolated rat atrial myocytes. Nicotine was applied at following concentrations: 4, 40 and 400 nM; ethanol at 20 mM (∼0.09%). Nicotine at 40 and 400 nM significantly activated constitutively active component of I K(Ach) with the maximum effect at 40 nM (an increase by ∼100%); similar effect was observed at -110 and -50 mV. Changes at 4 nM nicotine were negligible on average. Coapplication of 40 nM nicotine and 20 mM ethanol (which is also known to activate this current) did not show cumulative effect. In the case of acetylcholine-induced component of I K(Ach), a dual effect of nicotine and its correlation with the current magnitude in control were apparent: the current was increased by nicotine in the cells showing small current in control and vice versa. The effect of 40 and 400 nM nicotine on acetylcholine-induced component of I K(Ach) was significantly different at -110 and -50 mV. We conclude that nicotine at clinically relevant concentrations significantly increased constitutively active component of I K(Ach) and showed a dual effect on its acetylcholine-induced component, similarly as ethanol. Synchronous application of nicotine and ethanol did not cause additive effect.

• Klíčová slova
• Acetylcholine-sensitive, Dual effect, Inward rectifier, Nicotine,
• MeSH
• acetylcholin farmakologie   MeSH
• časové faktory MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny agonisté   účinky léků   MeSH
• ethanol toxicita   MeSH
• hodnocení rizik MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• membránové potenciály MeSH
• nikotin toxicita   MeSH
• potkani Wistar MeSH
• srdeční arytmie chemicky indukované   metabolismus   MeSH
• srdeční síně účinky léků   metabolismus   MeSH
• techniky in vitro MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetylcholin MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny MeSH
• ethanol MeSH
• nikotin MeSH

Antipsychotic drug perphenazine belongs to the phenothiazine group commonly reported to induce ECG changes and tachyarrhythmias. Data about its effect on ionic membrane currents in cardiomyocytes are missing. We analyzed the effect of perphenazine (0.1-100 microM) on fast sodium current I (Na) and transient outward potassium current I (to) in enzymatically isolated rat right ventricular myocytes by the whole-cell patch-clamp technique at room temperature. Perphenazine reversibly blocked I (Na) (reducing its amplitude; IC(50) = 1.24 +/- 0.10 microM) and I (to) (accelerating its apparent inactivation with a slight decrease of its amplitude; IC(50) = 38.2 +/- 3.5 microM, evaluated from changes of the time integral). The fast time constant of I (to) inactivation was significantly decreased in a concentration-dependent manner (IC(50) = 30.0 +/- 6.6 microM). Both blocks were use and frequency dependent at 3.3 Hz. We conclude that perphenazine causes concentration-, use-, and frequency-dependent block of I (Na) and I (to) . Computer simulations suggest that perphenazine interacts preferentially with I (Na) channels in inactivated states and with I (to) channels in both open and open-inactivated states.

• MeSH
• antipsychotika aplikace a dávkování   toxicita   MeSH
• draslíkové kanály účinky léků   metabolismus   MeSH
• inhibiční koncentrace 50 MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• krysa rodu Rattus MeSH
• metoda terčíkového zámku MeSH
• perfenazin aplikace a dávkování   toxicita   MeSH
• počítačová simulace MeSH
• potkani Wistar MeSH
• sodíkové kanály účinky léků   metabolismus   MeSH
• srdeční komory cytologie   účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antipsychotika MeSH
• draslíkové kanály MeSH
• perfenazin MeSH
• sodíkové kanály MeSH