Blebbistatin protects iPSC-CMs from hypercontraction and facilitates automated patch-clamp based electrophysiological...

Area of Science:

• Cardiovascular Research
• Electrophysiology
• Stem Cell Biology

Background:

• Human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) are crucial for cardiovascular channelopathy modeling and drug safety.
• Automated patch-clamp (APC) offers higher throughput than manual patch-clamp (MPC) but faces challenges with iPSC-CM quality.

Purpose of the Study:

• To optimize single iPSC-CM preparation for APC by improving cell quality during dissociation and recording.
• To assess the utility of APC with enhanced iPSC-CM preparation for modeling cardiac channelopathies.

Main Methods:

• Developed a modified APC protocol using 2.5 µM blebbistatin (BB) throughout iPSC-CM dissociation, filtration, storage, and recording.
• Compared iPSC-CM morphology and electrophysiological recordings (I_Na, spontaneous action potentials) with and without BB.
• Validated APC performance by comparing iPSC-CMs from Brugada syndrome patients with MPC data.

Main Results:

• Blebbistatin supplementation significantly improved iPSC-CM morphology and reduced blebbing in suspension.
• BB-treated iPSC-CMs yielded improved I_Na recordings and spontaneous action potentials.
• APC successfully recapitulated known electrophysiological phenotypes of Brugada syndrome patient-derived iPSC-CMs.

Conclusions:

• The optimized APC method using blebbistatin enhances iPSC-CM quality for electrophysiological studies.
• APC can effectively replace MPC for high-throughput modeling of cardiac channelopathies using iPSC-CMs.
• This approach facilitates advanced cardiovascular safety pharmacology and disease modeling.