Rapid, non-perfusion-limited calcium exchange in cultured neonatal myocardial cells

Neonatal rat myocardial cells culture are used to study the effects of zero [Ca]0 and lanthanum (La3+) on contractile function and to correlate these findings with steady-state 45Ca exchange. A rapid superfusion system is used to study cell shortening. At a stimulation rate of 1 Hz exposure to Ca2+ free solution for 900 ms abolishes contraction. Contraction is fully restored within 900 ms after return to 1 mM Ca2+ solution. Similarly, contraction is abolished 800 ms after changing to a 1 mM La3+ solution, but does not return for several min after removal of La3+. A modification of the scintillation-disc flow cell technique is used to study steady-state 45Ca exchange. Cells are grown directly on scintillant plastic discs pre-treated by the Primaria process which enhances cell adhesiveness. The cells on the discs are inserted into a specially designed flow cell which is then perfused at a rate of 500 ml/min. In this manner the labeling period, intervention and washout are performed while instantaneously and continuously monitoring the 45Ca activity without the need to interpose an unmonitored wash period. In addition, the use of on-line computer monitoring allows an accurate and precise recording of data as often as 6 times/s. This permits the identification of the following four Ca2+ compartments: (1) A rapidly exchanging compartment which is La3+ displaceable, contains 2.59 mmols Ca2+/kg dry cells and has a t1/2 = 1.5 s; (2) An intermediate component composed of two compartments which contain 0.43 and 0.22 mmols Ca2+/kg dry cells and exchange with t1/2 of 19 and 103 s respectively; (3) A slowly exchangeable compartment which contains 0.69 mmols Ca2+/kg dry cells with t1/2 = 19 min; (4) An unexchangeable compartment which contains 6.24 mmols Ca2+/kg dry cells. The rapid compartment is La3+ displaceable and has a flux of greater than 1100 mumols Ca2+/kg dry cells/s. This is the first time that the kinetics and sizes of the rapid Ca2+ compartment and intermediate compartments have been measured. Correlation of contractile response to zero [Ca]0 and La3+ administration with 45Ca exchange measurements under conditions of ultra-rapid perfusion indicate that at least a portion of the La3+-displaceable rapid component is critical in the beat-to-beat control of contractile force.