Mitochondria are involved in cell death for reasons that go beyond ATP supply. A recent advance has been the discovery that mitochondria contain and release proteins that are involved in the apoptotic cascade, like cytochrome c and apoptosis inducing factor. The involvement of mitochondria in cell death, and its being cause or consequence, remain issues that are extremely complex to address in situ. The response of mitochondria may critically depend on the type of stimulus, on its intensity, and on the specific mitochondrial function that has been primarily perturbed. On the other hand, the outcome also depends on the integration of mitochondrial responses that cannot be dissected easily. Here, we try to identify the mechanistic aspects of mitochondrial involvement in cell death as can be derived from our current understanding of mitochondrial physiology, with special emphasis on the permeability transition and its consequences (like onset of swelling, cytochrome c release and respiratory inhibition); and to critically evaluate methods that are widely used to monitor mitochondrial function in situ.

The opening of the mitochondrial permeability transition pore (PTP) has been suggested to play a key role in various forms of cell death, but direct evidence in intact tissues is still lacking. We found that in the rat heart, 92% of NAD(+) glycohydrolase activity is associated with mitochondria. This activity was not modified by the addition of Triton X-100, although it was abolished by mild treatment with the protease Nagarse, a condition that did not affect the energy-linked properties of mitochondria. The addition of Ca(2+) to isolated rat heart mitochondria resulted in a profound decrease in their NAD(+) content, which followed mitochondrial swelling. Cyclosporin A(CsA), a PTP inhibitor, completely prevented NAD(+) depletion but had no effect on the glycohydrolase activity. Thus, in isolated mitochondria PTP opening makes NAD(+) available for its enzymatic hydrolysis. Perfused rat hearts subjected to global ischemia for 30 min displayed a 30% decrease in tissue NAD(+) content, which was not modified by extending the duration of ischemia. Reperfusion resulted in a more severe reduction of both total and mitochondrial contents of NAD(+), which could be measured in the coronary effluent together with lactate dehydrogenase. The addition of 0.2 microm CsA or of its analogue MeVal-4-Cs (which does not inhibit calcineurin) maintained higher NAD(+) contents, especially in mitochondria, and significantly protected the heart from reperfusion damage, as shown by the reduction in lactate dehydrogenase release. Thus, upon reperfusion after prolonged ischemia, PTP opening in the heart can be documented as a CsA-sensitive release of NAD(+), which is then partly degraded by glycohydrolase and partly released when sarcolemmal integrity is compromised. These results demonstrate that PTP opening is a causative event in reperfusion damage of the heart.

The mitochondrial permeability transition pore is a high conductance channel whose opening leads to an increase of mitochondrial inner membrane permeability to solutes with molecular masses up to ≈ 1500 Da. In this review we trace the rise of the permeability transition pore from the status ofin vitroartifact to that of effector mechanism of cell death. We then cover recent results based on genetic inactivation of putative permeability transition pore components, and discuss their meaning for our understanding of pore structure. Finally, we discuss evidence indicating that the permeability transition pore plays a role in pathophysiology, with specific emphasis onin vivomodels of disease.

The mitochondrial permeability transition (PT) - an abrupt increase permeability of the inner membrane to solutes - is a causative event in ischemia-reperfusion injury of the heart, and the focus of intense research in cardioprotection. The PT is due to opening of the PT pore (PTP), a high conductance channel that is critically regulated by a variety of pathophysiological effectors. Very recent work indicates that the PTP forms from the F-ATP synthase, which would switch from an energy-conserving to an energy-dissipating device. This review provides an update on the current debate on how this transition is achieved, and on the PTP as a target for therapeutic intervention. This article is part of a Special Issue entitled "Mitochondria: from basic mitochondrial biology to cardiovascular disease".

Ischaemic postconditioning (brief periods of ischaemia alternating with brief periods of reflow applied at the onset of reperfusion following sustained ischaemia) effectively reduces myocardial infarct size in all species tested so far, including humans. Ischaemic postconditioning is a simple and safe manoeuvre, but because reperfusion injury is initiated within minutes of reflow, postconditioning must be applied at the onset of reperfusion. The mechanisms of protection by postconditioning include: formation and release of several autacoids and cytokines; maintained acidosis during early reperfusion; activation of protein kinases; preservation of mitochondrial function, most strikingly the attenuation of opening of the mitochondrial permeability transition pore (MPTP). Exogenous recruitment of some of the identified signalling steps can induce cardioprotection when applied at the time of reperfusion in animal experiments, but more recently cardioprotection was also observed in a proof-of-concept clinical trial. Indeed, studies in patients with an acute myocardial infarction showed a reduction of infarct size and improved left ventricular function when they underwent ischaemic postconditioning or pharmacological inhibition of MPTP opening during interventional reperfusion. Further animal studies and large-scale human studies are needed to determine whether patients with different co-morbidities and co-medications respond equally to protection by postconditioning. Also, our understanding of the underlying mechanisms must be improved to develop new therapeutic strategies to be applied at reperfusion with the ultimate aim of limiting the burden of ischaemic heart disease and potentially providing protection for other organs at risk of reperfusion injury, such as brain and kidney.