Fig 1.
Suggested voltage-clamp stimulation protocol and corresponding ionic currents recorded in human atrial myocytes.
A schematic representation of the voltage steps applied to elicit L-type calcium currents (ICa) and tail currents (ITail). A 50 ms prepulse from −80 to 45 mV is used to inactivate the sodium current (INa). Subsequently 200 ms depolarization from −45 to 0 mV elicits first a fast outward capacitive current (Icap) followed by and inward calcium current (ICa). Upon repolarization to –80 mV, a transient inward tail current (ITail) are recorded. This protocol was repeated for 30 consecutive pulses to assess beat-to-beat variability in calcium current responses.
Fig 2.
Features used for the alternating regime detection in patch-clamp calcium current recordings in the human atrial cardiomyocyte.
A. Pulsed voltage-clamp protocol applied to the cardiomyocyte across 30 sweeps. B–C. Representative consecutive current traces showing uniform and alternating beat-to-beat responses. The algorithm automatically identifies key features in each sweep, including: capacitive peak (PeakCap),calcium peak (PeakCa), tail peak (PeakTail), area of calcium current (QCa), area of tail current (QTail) and time constants (taus):tau (τ), tau 1 (τ₁), tau 2 (τ₂). D. Detailed view of alternating response. E. Detailed view of uniform response.
Fig 3.
Variation in calcium current area (QCa) and tail current area (Qtail) features in human atrial in cardiomyocytes with uniform and alternant electrical phenotype response.
A. Uniform response with stable current area (QCa) and tail current area (Qtail) amplitudes across 30 sweeps. B. Alternating response showing beat-to-beat alternations with current area (QCa) and tail current area (Qtail) out of phase. This relationship reflects the coupling between sarcolemmal calcium influx and sarcoplasmic reticulum (SR) calcium release.
Fig 4.
Electrical model of the whole-cell patch-clamp setup. A. Block diagram of a whole-cell voltage-clamp patch-clamp configuration using a single pipette. Electrical model of the whole-cell patch-clamp configuration used for parameter validation. B. The model combines a biophysical representation of the cardiomyocyte membrane and the recording hardware, including pipette resistance, amplifier feedback, and filtering circuits. This hybrid model generates realistic synthetic currents used to validate the feature extraction accuracy of the algorithm.
Table 1.
Electrical parameters used in the patch-clamp simulation using a zero-order model (electrical model). Simulation parameters have been obtained empirically from a real cardiomyocyte or through bibliography.
Fig 5.
Flowchart of the computational framework for characterizing experimental patch-clamp signals under pulsed clamp voltage.
The diagram starts by loading the data from the path definition, where the patch clamp event log files are stored. Then the current and voltage time series are extracted and properly pre-processed (filtering and interpolation). Then the current signal features are extracted. Finally, the data can be visualised, stored, or a basic statistic of the computed features can be computed.
Table 2.
AIC values for the single and double exponential models.
Fig 6.
Calculation of the alternation index in representative uniform and alternant cell regimes subjected to a stimulation protocol of 30 consecutive voltage pulses.
The variable D represents the difference between two consecutive values. A, B. Calculation of alternation index (CIdx) in a uniform and alternant cardiomyocyte response using calcium current area (QCa). C, D. Calculation of alternation index in a uniform and alternant cardiomyocyte response using tail current area (QTail).
Table 3.
Errors in automatically calculated features in a synthetic pulse.
Fig 7.
Comparison of peak amplitudes between uniform and alternant cardiomyocytes.
A. Amplitude of the L-type calcium current (ICa), is significantly lower (p ≤ 0.01) in alternant cells. B. Amplitude of the tail current (ITail) shows corresponding variability. These differences suggest impaired calcium influx and altered SR calcium handling in alternant cells.
Fig 8.
Time constant of ionic currents in uniform and alternating type cardiomyocytes.
A. Time constant of calcium current (τ). B. Time constant of the fast tail current (τ1). C. Time cost of the slow tail current (τ2). Alternant cells exhibit prolonged τ₁ (p ≤ 0.01) and shortened τ₂ (p ≤ 0.0001), indicating altered kinetics of calcium extrusion and SR reuptake.
Fig 9.
Areas under the current curves (AUC) for calcium and tail currents in uniform and alternant cardiomyocyte cells.
A. Area under the calcium current curve (QCa). This area is significantly smaller in alternant cardiomyocytes (p ≤ 0.001), reflecting reduced calcium influx. B. Area under the tail current curve (QTail). Shows no significant difference. Together, these data highlight the asymmetrical regulation of calcium entry and extrusion in alternant responses.
Fig 10.
Alternation index, CIdx, for uniform and alternating type cardiomyocytes derived from calcium current area (QCa) and the tail current area (QTail).
A. Alternation index calculated with the area under the calcium current curve (QCa). Differentiates alternating from uniform cells (p ≤ 0.01). B. Alternation index calculated with the area under the tail current curve (QTail), also shows significant discrimination (p ≤ 0.05).