Abstract
Liver mitochondria treated with N-ethylmaleimide can accumulate Ca2+ but cannot retain it. Ca2+ loss following uptake occurs in parallel with a proton uptake and collapse of the membrane potential. Respiration is not activated during Ca2+ release and cannot be stimulated by uncoupler. After Ca2+ release and accompanying phenomena are nearly complete, the mitochondria undergo a large amplitude swelling. Nupercaine inhibits the premature release of Ca2+, proton uptake, decline in membrane potential, inhibition of uncoupler-stimulated respiration, and large amplitude swelling. Ruthenium red also prevents these effects. Neither Sr2+ or Mn2+ will substitute for Ca2+ to induce these effects in N-ethylmaleimide-treated mitochondria. The effects of N-ethylmaleimide plus Ca2+ on mitochondria are not accompanied by a significant alteration in the content or composition of phospholipids but are accompanied by small increases in the mitochondrial content of free fatty acids. Free fatty acids accumulate more rapidly in response to limited Ca2+ loading in the absence of N-ethylmaleimide than they do in its presence. In the absence of N-ethylmaleimide, polyunsaturated fatty acids and saturated plus monounsaturated fatty acids accumulate at nearly equal rates. In the presence of N-ethylmaleimide, polyunsaturated fatty acids accumulate more rapidly than saturated plus monounsaturated fatty acids. Any condition or agent tested which inhibited swelling and the other effects produced by Ca2+ plus N-ethylmaleimide also prevented the more rapid accumulation of polyunsaturated, compared to saturated plus monounsaturated, fatty acids. In the light of a positional analysis of phospholipid acyl moieties, these data suggest that 1-acyllysophospholipids accumulate in swelling mitochondria but not in response to noraml Ca2+ loading or when swelling is blocked by other agents. The free fatty acid accumulation, per se, is not responsible for swelling, but levels of exogenous palmitic acid as low as 1 nmol/mg of protein dramatically alter the dependence of swelling velocity on Ca2+ concentration, producing a shift from a sigmoidal- to a hyperbolic-like relationship. This same alteration is brought about by aging the mitochondrial preparation at 0 degrees C. Either pyruvate or DL-carnitine prevents the effect of exogenous palmitate and restores the Aa2+ swelling dependence of aged N-ethylmaleimide-treated mitochondria to that of fresh N-ethylmaleimide-treated mitochondria. Intramitochondrial acylcoenzyme A or acylcarnitine, or both, therefore, to be the modulator of Ca2+ sensitivity rather than free fatty acid. The findings are discussed in terms of the role of intramitochondrial phospholipase and other phospholipid metabolizing enzymes in the mechanisms of N-ethylmaleimide plus Ca2+ effects on mitochondria.
Highlights
We proposed that the large amplitude swelling which is initiated by Ca” uptake into N-ethylmaleimide-treated mitochondria could result from an activation of a mitochondrial phospholipase A with resulting hydrolysis of inner mitochondrial membrane phospholipids and a concomitant increase in the permeability of the inner mitochondrial membrane [8]
As previously described [7, 8] and as seen in Fig. 1, Panel A, Ca2’ uptake by N-ethylmaleimide-treated mitochondria initiates the release of the Ca2+ with an accompanying H’ ion uptake and a large amplitude swelling of the mitochondria
The proton uptake, inhibition of uncoupler-stimulated respiration and decline in membrane potential occur on the same time course as CaZt movements rather than on the time course of swelling
Summary
Free fatty acids accumulate more rapidly in response to limited Ca2+ loading in the absence of N-ethylmaleimide than they do in its presence. In the absence of N-ethylmaleimide, polyunsaturated fatty acids and saturated plus monounsaturated fatty acids accumulate at nearly equal rates. In the presence of N-ethylmaleimide, polyunsaturated fatty acids accumulate more rapidly than saturated plus monounsaturated fatty acids. Any condition or agent tested which inhibited swelling and the other effects produced by Ca2+ plus N-ethylmaleimide prevented the more rapid accumulation of polyunsaturated, compared to saturated plus monounsaturated, fatty acids. The free fatty acid accumulation, per se, is not responsible for swelling, but levels of exogenous palmitic acid as low as 1 nmol/mg of protein dramatically alter the dependence of swelling velocity on Ca2+ concentration, producing a shift from a sigmoidal- to a hyperbolic-like relationship.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
