Journal: Membranes

Article Title: Evidence for a Physiological Role of T-Type Ca Channels in Ventricular Cardiomyocytes of Adult Mice

doi: 10.3390/membranes12060566

Figure Lengend Snippet: Ca currents in wt and Cav3.2 KO ventricular cardiomyocytes. ( A ) Original traces of Ca currents of a typical wt and Cav3.2 KO myocyte elicited by the pulse protocol displayed on top. ( B ) Current density–voltage relationships of wt (n = 20) and Cav3.2 KO (n = 20) myocytes. Data are expressed as means ± SEM. The current density–voltage relations hardly revealed any T-type Ca channel activity (expected to be maximal at −45 mV) but did reveal significant L-type Ca channel activity (peaking at +5 mV), which was similar in wt and Cav3.2 KO myocytes ( p = 0.67 at +5 mV, unpaired Student’s t -test). Capacitance values of the examined myocytes were 184 ± 13 pF for wt and 169 ± 11 pF for Cav3.2 KO ( p = 0.37). ( C ) The effect of the external application of 100 nM isoprenaline (ISO) on ICaT in a wt myocyte. From a holding potential of −95 mV, depolarizing voltage steps to −45 mV were applied every 3 s to elicit the currents. The arrows indicate the beginning and end of ISO application. Current peaks were finally plotted against time. ( D ) ICaT peaks before, during application after the steady-state was reached, and after the washout of ISO in wt (top) and Cav3.2 KO (bottom) myocytes. There was no significant difference between wt and Cav3.2 KO under control conditions ( p = 0.81, unpaired Student’s t -test) or in the presence of ISO ( p = 0.54). Data are expressed as means ± SEM. Each data point represents a single cell. ** p < 0.01; *** p < 0.001; paired Student’s t -test.

Article Snippet: This was followed by blocking with 10% goat serum (Sigma-Aldrich, Vienna, Austria) and 0.01% azide (Sigma-Aldrich, Vienna, Austria) in PBS for 2 h. Subsequently, the cells were incubated with a selective anti-Cav3.1 antibody (anti-CACNA1G, #ACC-021, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:500) or with one of two selective anti-Cav3.2 antibodies (anti-CACNA1H, #ACC-025, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:1000; anti-Cav3.2 polyclonal antibody, #PA5-106771, source: rabbit, Invitrogen, Paisley, England, 1:200) at 4 °C overnight.

Techniques: Activity Assay

Journal: Membranes

Article Title: Evidence for a Physiological Role of T-Type Ca Channels in Ventricular Cardiomyocytes of Adult Mice

doi: 10.3390/membranes12060566

Figure Lengend Snippet: Immunostaining of T-type Ca channels in ventricular cardiomyocytes isolated from adult wt mice. Cav3.1 ( top ) channel expression and localization was detected using a selective anti-Cav3.1 antibody (anti-CACNA1G, #ACC-021, source: rabbit, Alomone Labs, 1:500). Cav3.2 ( bottom ) was detected with the selective anti-Cav3.2 antibodies (anti-CACNA1H, #ACC-025, source: rabbit, Alomone Labs, 1:1000; left and right cell; or anti-Cav3.2 polyclonal, #PA5-106771, source: rabbit, Invitrogen, 1:200; middle cell). Secondary antibody: Alexa Fluor 488, #A11008, goat anti-rabbit, Invitrogen, 1:500.

Article Snippet: This was followed by blocking with 10% goat serum (Sigma-Aldrich, Vienna, Austria) and 0.01% azide (Sigma-Aldrich, Vienna, Austria) in PBS for 2 h. Subsequently, the cells were incubated with a selective anti-Cav3.1 antibody (anti-CACNA1G, #ACC-021, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:500) or with one of two selective anti-Cav3.2 antibodies (anti-CACNA1H, #ACC-025, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:1000; anti-Cav3.2 polyclonal antibody, #PA5-106771, source: rabbit, Invitrogen, Paisley, England, 1:200) at 4 °C overnight.

Techniques: Immunostaining, Isolation, Expressing

Journal: Membranes

Article Title: Evidence for a Physiological Role of T-Type Ca Channels in Ventricular Cardiomyocytes of Adult Mice

doi: 10.3390/membranes12060566

Figure Lengend Snippet: Electrically induced and caffeine-induced intracellular Ca transients in wt and Cav3.2 KO ventricular cardiomyocytes. ( A ) Left: Representative time courses of Fluo-4 fluorescence reporting rises in cytosolic Ca concentration during electrical field stimulation at 0.1 Hz frequency in a single wt and Cav3.2 KO myocyte. The orange lines represent fits of the data with a single exponential function. Right: Comparison of mean Ca peak fluorescence relative to baseline (F/F0) between wt and Cav3.2 KO myocytes. Each data point represents a single cell, and values are expressed as means ± SEM (n= 68 for wt and 59 for Cav3.2 KO myocytes). * p < 0.05 (0.02); unpaired Student’s t -test. The Ca transients decayed with time constants of 0.30 ± 0.02 s in wt and 0.25 ± 0.01 s in Cav3.2 KO myocytes, which were significantly different ( p = 0.04). ( B ) Comparison of mean Ca peak fluorescence (F/F0) before and after the application of 100 nM ISO in wt (n = 20) and Cav3.2 KO (n = 21) myocytes. **** p < 0.0001 (paired Student’s t -test). There was no significant difference between wt and Cav3.2 KO in the presence of ISO ( p = 0.37; unpaired Student’s t -test). The Ca transients under ISO decayed with time constants of 0.14 ± 0.02 s in wt and 0.11 ± 0.02 s in Cav3.2 KO myocytes (not significantly different, p = 0.19). ( C ) Comparison of mean Ca peak fluorescence elicited by an application of 20 mM caffeine, relative to baseline (F/F0), between wt (n = 26) and Cav3.2 KO (n = 35) myocytes. ns—not significant (unpaired Student’s t -test, p = 0.42).

Article Snippet: This was followed by blocking with 10% goat serum (Sigma-Aldrich, Vienna, Austria) and 0.01% azide (Sigma-Aldrich, Vienna, Austria) in PBS for 2 h. Subsequently, the cells were incubated with a selective anti-Cav3.1 antibody (anti-CACNA1G, #ACC-021, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:500) or with one of two selective anti-Cav3.2 antibodies (anti-CACNA1H, #ACC-025, source: rabbit, Alomone Labs, Jerusalem, Israel, 1:1000; anti-Cav3.2 polyclonal antibody, #PA5-106771, source: rabbit, Invitrogen, Paisley, England, 1:200) at 4 °C overnight.

Techniques: Fluorescence, Concentration Assay

Journal: MethodsX

Article Title: Multipurpose setup used to characterize tribo-electrical properties of electrical contact materials

doi: 10.1016/j.mex.2021.101498

Figure Lengend Snippet: Components required.

Article Snippet: Independently, the setup is equipped with a current/voltage sourcing unit (Keithley 2400 SMU) and a nanovoltmeter (Keithley 2182a).

Techniques:

Journal: MethodsX

Article Title: Multipurpose setup used to characterize tribo-electrical properties of electrical contact materials

doi: 10.1016/j.mex.2021.101498

Figure Lengend Snippet: Calculated resistance standard deviation for different voltage ranges.

Article Snippet: Independently, the setup is equipped with a current/voltage sourcing unit (Keithley 2400 SMU) and a nanovoltmeter (Keithley 2182a).

Techniques: Standard Deviation

Journal: MethodsX

Article Title: Multipurpose setup used to characterize tribo-electrical properties of electrical contact materials

doi: 10.1016/j.mex.2021.101498

Figure Lengend Snippet:

Article Snippet: Independently, the setup is equipped with a current/voltage sourcing unit (Keithley 2400 SMU) and a nanovoltmeter (Keithley 2182a).

Techniques: