What Is Myocardial Action Potential?

Cardiac cells, also known as myocardial fibers, have stripes and are dominated by vegetative nerves. They belong to involuntary muscles with stripes and have the ability to excite and contract. It has a short cylindrical shape with branches, and its nucleus is located in the center of the cell, usually only one. The ends of each myocardial fiber branch can be connected to each other to form a muscle fiber network. Broadly defined myocardial cells include the specially differentiated myocardial cells that make up the sinoatrial node, intraatrial bundle, atrioventricular junction, atrioventricular bundle (ie, Heath bundle), and Purkinje fibers, as well as general atrial and ventricular muscles. Working cells.

According to their

1. Myocardial cells are short columnar, generally have only one nucleus, while skeletal muscle fibers are

Cardiomyocytes

Cardiomyocyte hypertrophy

Myocardial tissue includes myocardial cells and

Ion channels and ion currents of cardiomyocytes are the main content of electrophysiology of cardiomyocytes. Its research progress is very rapid, especially the combination of molecular biology research, it can be said that it has made rapid progress. Nowadays, the electrophysiology of cardiac muscle cells, especially

Culture and isolation of rat cardiomyocytes

purpose

A cardiomyocyte culture model was established for cardiomyocyte in vitro experiments.

method

The myocardial cells were isolated and purified by trypsin digestion and the differential adherence method between neonatal rat cardiomyocytes and non-cardiomyocytes such as fibroblasts and blood cells to establish a myocardial culture model.

result

The longest survival time of cardiomyocytes is 3 days, and the cardiomyocytes have different morphological effects due to the effect of digestion time.

in conclusion

Neonatal rat cardiomyocytes can be isolated and cultured in vitro using the time-adherent method. Low-concentration and short-term trypsin digestion is preferred.

material

Digestion of cardiomyocytes

Take a covered beaker, place a piece of gauze saturated with ether, place 0 to 3 days old rat pups in the beaker and anaesthetize, disinfect the chest and abdomen skin with 2% iodine and 75% alcohol. The heart was removed under thoracotomy conditions, and immediately placed in 4 & ordm; CD-Hanks solution (mmol / L: Nacl137, Kcl5.4, Na2HPO40.37, K2HPO40.44, NaHCO34.2) to cut the ventricular muscles, and wash the residual blood , Cut into tissue pieces of about 1mm & sup; size, discard D-Hanks solution and add 10 ~ 15ml of 0.08% trypsin solution, leave it at 37 ° C for 5min, aspirate the upper suspension, and add the same amount of serum-containing medium. After termination of digestion, the supernatant was discarded by centrifugation (1800 r / min), and a serum-containing culture solution was added. Blow down the pelleted cells, centrifuge under the same conditions, make a cell suspension with 10% serum culture solution, and place in a 37 ° C incubator containing 5% CO2.

Cardiomyocyte isolation

Two-hour differential adherence was used to obtain myocardial fibroblasts and cardiomyocytes according to the different attachment times of cardiomyocytes and non-cardiac cells. Cardiomyocytes were seeded in 50 ml of culture medium at 1 * 106 cells / ml, and 5-bromodeoxypyrimidine nucleoside 0.1 mmol / l was added to the myocardial cell culture medium 2 days before the culture to inhibit the proliferation of non-cardiomyocytes. Myocardial cells were changed every 2 days.

1.3 Serum-free culture of cardiomyocytes

When the cardiomyocytes are cultured for 24 hours, change the serum-free medium (containing DMEM medium, insulin 10g / ml, ferritin 10g / ml, vitamin C100g / L, vitamin B121.5 & micro; mol / L) and continue to culture For 48 hours, change the solution every 8 hours, try to keep the concentration of each added component unchanged, and finally collect cells for measurement.

2. Experimental results

In this experiment, the longest survival time of neonatal rat cardiomyocyte culture was 3 days. The experiment adopts low concentration and short time trypsin digestion method. Different digestion time, the resulting cells have different morphological changes: 3 to 5 minutes of digestion, the majority of myocardial cells were observed at high magnification to be crescent-shaped, and it can be seen that the cells show a head-to-tail connection process in the culture medium, showing dynamic changes . The cardiomyocytes that are digested for 5 to 10 minutes are round and have low density and poor mobility. After about 8 hours of cell culture, the cells gathered in one place and hooked to each other to show a high-density area, which may be related to the formation of connections between cells. For some unknown reason, unknown microorganisms appeared in myocardial cells during multiple cultures, causing the cells to basically die after 3 days.

3. Experimental discussion

Many literatures have reported that the culture time of neonatal rat cardiomyocytes is 10-12 days, and the culture time of this experiment is 3 days. Compared with this, the survival time is shorter. The digestion time is better than the general concentration (0.125%). The digestion time can reduce the cell death rate within 3 ~ 5 minutes. The phenomenon of cells gathering in one place and the visible continuous morphological changes during the experiment can explain It is possible to reconnect myocardial cells in vitro to form larger unit cell clusters, which may be linked to each other through network changes.