(26) RAPID EFFLUX OF Ca2+ FROM LIVER AND HEART MITOCHONDRIA

K. Gunter, L. Buntinas, R. Eliseev, G. Sparagna and T. Gunter
Department of Biochemistry and Biophysics, University of Rochester, Rochester, NY

Rapid efflux of Ca2+ from liver and heart mitochondria can be induced following Ca2+ uptake via RaM by dropping the external [Ca2+] to a low level. It was observed after exposing mitochondria to pulses of Ca2+ generated by our pulse-generating system, consisting of a computer-controlled automatic pipetter used in conjunction with a fluorescence spectrometer.

When two mitochondrial aliquots are treated identically with respect to [Ca2+] before and during the pulse and sequester Ca2+ via the RaM mechanism, using different [Ca2+]’s at the end of the pulse can lead to different Ca2+ retention. If the [Ca2+] following the pulse for the first aliquot is dropped to 100 nM while the [Ca2+] following the pulse for the second aliquot is 20 nM, considerably more Ca2+ will be retained by the mitochondria in the first aliquot. Raising the [Ca2+] bathing the second aliquot to 100 nM within seconds of its fall to 20 nM and before separating the mitochondria from their supernatant by centrifugation does nothing to restore the Ca2+ retained by that aliquot. Since the Ca2+ history of the aliquots is identical prior to addition of differing amounts of a chelator, such as EGTA, to end the pulse, the observed differences in retained Ca2+ must be due to efflux of some Ca2+ following chelator addition. The ‘threshold” for Ca2+ loss of this type seems to be approximately the same as the kd for Ca2+ binding to the external inhibitor site of the RaM.

RaM uptake of Ca2+,like that of the uniporter can be eliminated by deenergizing the mitochondria. Since movement of Ca2+ outward across the large mitochondrial membrane potential is energetically uphill, loss of Ca2+ through this rapid efflux must involve some form of at least local deenergization of the mitochondria.

Other characteristics of this phenomenon will be discussed.

Supported by AHA NY State Affiliate Grant # 960166.