Research

Abstract: The aim of this study was to use both a theoretical and experimental approach to determine the influence of the sarco-endoplasmic Ca2+-ATPase (SERCA) activity and mitochondria Ca2+ uptake on Ca2+ homeostasis in airway myocytes. Experimental studies were performed on myocytes freshly isolated from rat trachea. [Ca2+]i was measured by microspectrofluorimetry using indo-1. Stimulation by caffeine for 30 s induced a concentration-graded response characterized by a transient peak followed by a progressive decay to a plateau phase. The decay phase was accelerated for 1-s stimulation, indicating ryanodine receptor closure. In Na2+-Ca2+-free medium containing 0.5 mM La3+, the [Ca2+]i response pattern was not modified, indicating no involvement of transplasmalemmal Ca2+ fluxes. The mathematical model describing the mechanism of Ca2+ handling upon RyR stimulation predicts that after Ca2+ release from the sarcoplasmic reticulum, the Ca2+ is first sequestrated by cytosolic proteins and mitochondria, and pumped back into the sarcoplasmic reticulum after a time delay. Experimentally, we showed that the [Ca2+]i decay after Ca2+ increase was not altered by the SERCA inhibitor cyclopiazonic acid, but was slightly but significantly modified by the mitochondria uncoupler carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone. The experimental and theoretical results indicate that, although Ca2+ pumping back by SERCA is active, it is not primarily involved in [Ca2+]i decrease that is due, in part, to mitochondrial Ca2+ uptake.

The Biophysical Society	*E. Roux and M. Marhl* Role of sarcoplasmic reticulum and mitochondria in Ca2+ removal in airway myocytes Biophysical Journal 86 (2004) 2583-2595	Abstract: The aim of this study was to use both a theoretical and experimental approach to determine the influence of the sarco-endoplasmic Ca2+-ATPase (SERCA) activity and mitochondria Ca2+ uptake on Ca2+ homeostasis in airway myocytes. Experimental studies were performed on myocytes freshly isolated from rat trachea. [Ca2+]i was measured by microspectrofluorimetry using indo-1. Stimulation by caffeine for 30 s induced a concentration-graded response characterized by a transient peak followed by a progressive decay to a plateau phase. The decay phase was accelerated for 1-s stimulation, indicating ryanodine receptor closure. In Na2+-Ca2+-free medium containing 0.5 mM La3+, the [Ca2+]i response pattern was not modified, indicating no involvement of transplasmalemmal Ca2+ fluxes. The mathematical model describing the mechanism of Ca2+ handling upon RyR stimulation predicts that after Ca2+ release from the sarcoplasmic reticulum, the Ca2+ is first sequestrated by cytosolic proteins and mitochondria, and pumped back into the sarcoplasmic reticulum after a time delay. Experimentally, we showed that the [Ca2+]i decay after Ca2+ increase was not altered by the SERCA inhibitor cyclopiazonic acid, but was slightly but significantly modified by the mitochondria uncoupler carbonyl cyanide 4-(trifluoromethoxy) phenylhydrazone. The experimental and theoretical results indicate that, although Ca2+ pumping back by SERCA is active, it is not primarily involved in [Ca2+]i decrease that is due, in part, to mitochondrial Ca2+ uptake.

Elsevier	*V. Grubelnik, A.Z. Larsen, U. Kummer, L.F. Olsen, M. Marhl* Mitochondria regulate the amplitude of simple and complex calcium oscillations Biophysical Chemistry 94 (2001) 59-74	Abstract: In a mathematical model for simple calcium oscillations [Biophys. Chem. 71 (1998) 125], it has been shown that mitochondria play an important role in the maintenance of constant amplitudes of cytosolic Calf oscillations. Simple plausible rate laws for Calf fluxes across the inner mitochondrial membrane have been used in this model. Here we show that it is possible to use the same rate laws as a plug-in element in other existing mathematical models and obtain the same effect on amplitude regulation. This result appears to be universal, independent of the type of model and the type of Ca2+ oscillations. We demonstrate this on two models for spiking Calf oscillations [J. Biol. Chem. 266 (1991) 11068; Cell Calcium 14 (1993) 311] and on two recent models for bursting Ca2+ oscillations; one of them being a receptor-operated model [Biophys. J. 79 (2000) 1188] and the other one being a store-operated model [BioSystems 57 (2000) 75].

Elsevier	*M. Marhl, T. Haberichter, M. Brumen, R. Heinrich* Complex calcium oscillations and the role of mitochondria and cytosolic proteins Biosystems 57 (2000) 75-86	Abstract: Intracellular calcium oscillations, which are oscillatory changes of cytosolic calcium concentration in response to agonist stimulation, are experimentally well observed in various living cells. Simple calcium oscillations represent the most common pattern and many mathematical models have been published to describe this type of oscillation. On the other hand, relatively few theoretical studies have been proposed to give an explanation of complex intracellular calcium oscillations, such as bursting and chaos. In this paper, we develop a new possible mechanism for complex calcium oscillations based on the interplay between three calcium stores in the cell: the endoplasmic reticulum (ER), mitochondria and cytosolic proteins. The majority (approximate to 80%) of calcium released from the ER is first very quickly sequestered by mitochondria. Afterwards, a much slower release of calcium from the mitochondria serves as the calcium supply for the intermediate calcium exchanges between the ER and the cytosolic proteins causing bursting calcium oscillations. Depending on the permeability of the ER channels and on the kinetic properties of calcium binding to the cytosolic proteins, different patterns of complex calcium oscillations appear. With our model, we are able to explain simple calcium oscillations, bursting and chaos. Chaos is also observed for calcium oscillations in the bursting mode.

Elsevier	*M. Marhl, S. Schuster, M. Brumen* Mitochondria as an important factor in the maintenance of constant amplitudes of cytosolic calcium oscillations Biophysical Chemistry 71 (1998) 125-132	Abstract: Theoretical models of intracellular calcium oscillations have hitherto focused on the endoplasmic reticulum (ER) as an internal calcium store. These models reproduced the large variability in oscillation frequency observed experimentally. In the present contribution, we extend our earlier model [Marhl et al., Biophys, Chem., 63 (1997) 221] by including, in addition to the ER, mitochondria as calcium stores. Simple plausible rate laws are used for the calcium uptake into, and release from, the mitochondria. It is demonstrated with the help of this extended model that mitochondria are likely to act in favour of frequency encoding by enabling the maintenance of fairly constant amplitudes over wide ranges of frequency.