Until recently, the rate of ATP production was thought to be determined by the rate at which ADP and phosphate (Pi) diffuse back to mitochondria. Recent evidence at the cellular and tissue levels suggests control by a novel mechanism, probably functioning through intramitochondrial [Ca2+]. 31P NMR has identified conditions in which [ADP] and [Pi] remain constant while ATP production is increased by a factor of four or more. Clearly, metabolic rate cannot be activated by increased [ADP] and [Pi] if they do not increase, and another mechanism of control is indicated. This additional mechanism is thought to involve intramitochondrial free calcium ([Ca2+]m). Therefore, it is important to determine whether enough Ca2+ can be sequestered by mitochondria under physiological conditions to serve this function of metabolic mediator (1).

Under physiological conditions, cytosolic free calcium ([Ca2+]c) in many tissues remains low (80 to 100 nM) except during pulses or transients of [Ca2+]c. During these pulses, [Ca2+]c can become 1 µM or larger. Even liver, a non excitable tissue, may respond to hormones through a sequence of Ca2+ pulses. A typical hepatocyte response to vasopressin, for example, could be a sequence of 6 or 8 Ca2+ pulses. It is important to determine if mitochondria can sequester enough Ca2+ from such pulses to activate the Ca2+-sensitive steps of the metabolic pathways (1). Calculations based on the kinetics of known mitochondrial Ca2+ transporters suggested that they cannot sequester enough Ca2+(1). However, these kinetics were determined using buffered [Ca2+], not [Ca2+] pulses as under physiological conditions.

We built a device capable of generating Ca2+ pulses like those observed in vivo in many tissues. The [Ca2+] is controlled by a computer-controlled automatic pipetter and measured using fluorescence. We can generate [Ca2+] pulses down to durations of 0.2 - 0.3 sec. over a broad range of [Ca2+]. Using this device, we have discovered a new mechanism of Ca2+ uptake into liver mitochondria, termed the RaM ("rapid mechanism"). Controls show that the RaM mediates rapid net mitochondrial uptake from Ca2+ pulses (2). The RaM briefly displays very high Ca2+ conductivity at the beginning of a pulse; however, the RaM is rapidly closed as the [Ca2+] of the pulse increases. It is quickly "reset" by the fall in [Ca2+] between Ca2+ pulses and therefore functions at the beginning of each pulse. It is sufficiently activated by physiological concentrations of spermine to allow enough Ca2+ to be sequestered from a few pulses to stimulate ATP production. RaM-mediated metabolic signaling shows characteristics of "frequency modulation" (2). The RaM also exists in heart mitochondria; however, its characteristics in heart are quite different from those observed in liver. We believe that this newly discovered mechanism may be the most important component of the system controlling metabolic rate.

• Gunter TE, Gerstner B, Gunter KK, Malecki J, Gelein R, Valentine WM, Aschner M, Yule DI. Manganese transport via the transferrin mechanism. Neurotoxicology. 2013 Jan; 34:118-27.
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• Gunter TE, Gerstner B, Lester T, Wojtovich AP, Malecki J, Swarts SG, Brookes PS, Gavin CE, Gunter KK. An analysis of the effects of Mn2+ on oxidative phosphorylation in liver, brain, and heart mitochondria using state 3 oxidation rate assays. Toxicol Appl Pharmacol. 2010 Nov 15; 249(1):65-75.
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• Gunter TE, Gavin CE, Gunter KK. The case for manganese interaction with mitochondria. Neurotoxicology. 2009 Jul; 30(4):727-9.
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• Eliseev RA, Malecki J, Lester T, Zhang Y, Humphrey J, Gunter TE. Cyclophilin D interacts with Bcl2 and exerts an anti-apoptotic effect. J Biol Chem. 2009 Apr 10; 284(15):9692-9.
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• Gunter TE, Sheu SS. Characteristics and possible functions of mitochondrial Ca(2+) transport mechanisms. Biochim Biophys Acta. 2009 Nov; 1787(11):1291-308.
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• Brookes PS, Parker N, Buckingham JA, Vidal-Puig A, Halestrap AP, Gunter TE, Nicholls DG, Bernardi P, Lemasters JJ, Brand MD. UCPs--unlikely calcium porters. Nat Cell Biol. 2008 Nov; 10(11):1235-7; author reply 1237-40.
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• Eliseev RA, Filippov G, Velos J, VanWinkle B, Goldman A, Rosier RN, Gunter TE. Role of cyclophilin D in the resistance of brain mitochondria to the permeability transition. Neurobiol Aging. 2007 Oct; 28(10):1532-42.
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• Gunter TE, Gavin CE, Aschner M, Gunter KK. Speciation of manganese in cells and mitochondria: a search for the proximal cause of manganese neurotoxicity. Neurotoxicology. 2006 Sep; 27(5):765-76.
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• Gunter KK, Aschner M, Miller LM, Eliseev R, Salter J, Anderson K, Gunter TE. Determining the oxidation states of manganese in NT2 cells and cultured astrocytes. Neurobiol Aging. 2006 Dec; 27(12):1816-26.
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• Gunter KK, Aschner M, Miller LM, Eliseev R, Salter J, Anderson K, Hammond S, Gunter TE. Determining the oxidation states of manganese in PC12 and nerve growth factor-induced PC12 cells. Free Radic Biol Med. 2005 Jul 15; 39(2):164-81.
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• Eliseev RA, Vanwinkle B, Rosier RN, Gunter TE. Diazoxide-mediated preconditioning against apoptosis involves activation of cAMP-response element-binding protein (CREB) and NFkappaB. J Biol Chem. 2004 Nov 5; 279(45):46748-54.
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• Gunter TE, Yule DI, Gunter KK, Eliseev RA, Salter JD. Calcium and mitochondria. FEBS Lett. 2004 Jun 1; 567(1):96-102.
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• Eliseev R, Alexandrov A, Gunter T. High-yield expression and purification of p18 form of Bax as an MBP-fusion protein. Protein Expr Purif. 2004 Jun; 35(2):206-9.
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• Gunter TE, Miller LM, Gavin CE, Eliseev R, Salter J, Buntinas L, Alexandrov A, Hammond S, Gunter KK. Determination of the oxidation states of manganese in brain, liver, and heart mitochondria. J Neurochem. 2004 Jan; 88(2):266-80.
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• Eliseev RA, Gunter KK, Gunter TE. Bcl-2 prevents abnormal mitochondrial proliferation during etoposide-induced apoptosis. Exp Cell Res. 2003 Oct 1; 289(2):275-81.
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• Eliseev RA, Salter JD, Gunter KK, Gunter TE. Bcl-2 and tBid proteins counter-regulate mitochondrial potassium transport. Biochim Biophys Acta. 2003 Apr 18; 1604(1):1-5.
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• Zuscik MJ, O'Keefe RJ, Gunter TE, Puzas JE, Schwarz EM, Rosier RN. Parathyroid hormone-related peptide regulation of chick tibial growth plate chondrocyte maturation requires protein kinase A. J Orthop Res. 2002 Sep; 20(5):1079-90.
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• Zuscik MJ, D'Souza M, Ionescu AM, Gunter KK, Gunter TE, O'Keefe RJ, Schwarz EM, Puzas JE, Rosier RN. Growth plate chondrocyte maturation is regulated by basal intracellular calcium. Exp Cell Res. 2002 Jun 10; 276(2):310-9.
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• Eliseev RA, Gunter KK, Gunter TE. Bcl-2 sensitive mitochondrial potassium accumulation and swelling in apoptosis. Mitochondrion. 2002 Feb; 1(4):361-70.
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• Pfeiffer DR, Gunter TE, Eliseev R, Broekemeier KM, Gunter KK. Release of Ca2+ from mitochondria via the saturable mechanisms and the permeability transition. IUBMB Life. 2001 Sep-Nov; 52(3-5):205-12.
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• Gunter TE, Gunter KK. Uptake of calcium by mitochondria: transport and possible function. IUBMB Life. 2001 Sep-Nov; 52(3-5):197-204.
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• Gunter KK, Gunter TE. Measurements of intracellular free calcium concentration in biological systems. Curr Protoc Toxicol. 2001 May; Chapter 2:Unit2.5.
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• Buntinas L, Gunter KK, Sparagna GC, Gunter TE. The rapid mode of calcium uptake into heart mitochondria (RaM): comparison to RaM in liver mitochondria. Biochim Biophys Acta. 2001 Apr 2; 1504(2-3):248-61.
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• Gunter TE, Buntinas L, Sparagna G, Eliseev R, Gunter K. Mitochondrial calcium transport: mechanisms and functions. Cell Calcium. 2000 Nov-Dec; 28(5-6):285-96.
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• Gavin CE, Gunter KK, Gunter TE. Manganese and calcium transport in mitochondria: implications for manganese toxicity. Neurotoxicology. 1999 Apr-Jun; 20(2-3):445-53.
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• Gunter TE, Buntinas L, Sparagna GC, Gunter KK. The Ca2+ transport mechanisms of mitochondria and Ca2+ uptake from physiological-type Ca2+ transients. Biochim Biophys Acta. 1998 Aug 10; 1366(1-2):5-15.
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• Gunter TE, Buntinas L, Sparagna GC, Gunter KK. The interaction of mitochondria with pulses of calcium. Biofactors. 1998; 8(3-4):205-7.
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• Zuscik MJ, Gunter TE, Puzas JE, Rosier RN. Characterization of voltage-sensitive calcium channels in growth plate chondrocytes. Biochem Biophys Res Commun. 1997 May 19; 234(2):432-8.
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• Sparagna GC, Gunter KK, Sheu SS, Gunter TE. Mitochondrial calcium uptake from physiological-type pulses of calcium. A description of the rapid uptake mode. J Biol Chem. 1995 Nov 17; 270(46):27510-5.
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• Zuscik MJ, Puzas JE, Rosier RN, Gunter KK, Gunter TE. Cyclic-AMP-dependent protein kinase activity is not required by parathyroid hormone to stimulate phosphoinositide signaling in chondrocytes but is required to transduce the hormone's proliferative effect. Arch Biochem Biophys. 1994 Dec; 315(2):352-61.
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• Gunter TE. Cation transport by mitochondria. J Bioenerg Biomembr. 1994 Oct; 26(5):465-9.
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• Gunter KK, Gunter TE. Transport of calcium by mitochondria. J Bioenerg Biomembr. 1994 Oct; 26(5):471-85.
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• Zuscik MJ, Gunter TE, Rosier RN, Gunter KK, Puzas JE. Activation of phosphoinositide metabolism by parathyroid hormone in growth plate chondrocytes. Cell Calcium. 1994 Aug; 16(2):112-22.
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• Gunter TE, Gunter KK, Sheu SS, Gavin CE. Mitochondrial calcium transport: physiological and pathological relevance. Am J Physiol. 1994 Aug; 267(2 Pt 1):C313-39.
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• Sparagna GC, Gunter KK, Gunter TE. A system for producing and monitoring in vitro calcium pulses similar to those observed in vivo. Anal Biochem. 1994 May 15; 219(1):96-103.
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• Baysal K, Jung DW, Gunter KK, Gunter TE, Brierley GP. Na(+)-dependent Ca2+ efflux mechanism of heart mitochondria is not a passive Ca2+/2Na+ exchanger. Am J Physiol. 1994 Mar; 266(3 Pt 1):C800-8.
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• Gavin CE, Gunter KK, Gunter TE. Mn2+ sequestration by mitochondria and inhibition of oxidative phosphorylation. Toxicol Appl Pharmacol. 1992 Jul; 115(1):1-5.
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• Gunter KK, Zuscik MJ, Gunter TE. The Na(+)-independent Ca2+ efflux mechanism of liver mitochondria is not a passive Ca2+/2H+ exchanger. J Biol Chem. 1991 Nov 15; 266(32):21640-8.
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• Gavin CE, Gunter KK, Gunter TE. Mn2+ transport across biological membranes may be monitored spectroscopically using the Ca2+ indicator dye antipyrylazo III. Anal Biochem. 1991 Jan; 192(1):44-8.
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• Gunter TE, Zuscik MJ, Puzas JE, Gunter KK, Rosier RN. Cytosolic free calcium concentrations in avian growth plate chondrocytes. Cell Calcium. 1990 Aug; 11(7):445-57.
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• Gunter TE, Pfeiffer DR. Mechanisms by which mitochondria transport calcium. Am J Physiol. 1990 May; 258(5 Pt 1):C755-86.
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• Gavin CE, Gunter KK, Gunter TE. Manganese and calcium efflux kinetics in brain mitochondria. Relevance to manganese toxicity. Biochem J. 1990 Mar 1; 266(2):329-34.
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• Gunter TE, Restrepo D, Gunter KK. Conversion of esterified fura-2 and indo-1 to Ca2+-sensitive forms by mitochondria. Am J Physiol. 1988 Sep; 255(3 Pt 1):C304-10.
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• Gunter TE, Wingrove DE, Banerjee S, Gunter KK. Mechanisms of mitochondrial calcium transport. Adv Exp Med Biol. 1988; 232:1-14.
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• Wingrove DE, Gunter TE. Kinetics of mitochondrial calcium transport. II. A kinetic description of the sodium-dependent calcium efflux mechanism of liver mitochondria and inhibition by ruthenium red and by tetraphenylphosphonium. J Biol Chem. 1986 Nov 15; 261(32):15166-71.
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• Wingrove DE, Gunter TE. Kinetics of mitochondrial calcium transport. I. Characteristics of the sodium-independent calcium efflux mechanism of liver mitochondria. J Biol Chem. 1986 Nov 15; 261(32):15159-65.
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• Gunter TE, Jensen BD. The efficiencies of the component steps of oxidative phosphorylation. I. A simple steady state theory. Arch Biochem Biophys. 1986 Jul; 248(1):289-304.
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• Jensen BD, Gunter KK, Gunter TE. The efficiencies of the component steps of oxidative phosphorylation. II. Experimental determination of the efficiencies in mitochondria and examination of the equivalence of membrane potential and pH gradient in phosphorylation. Arch Biochem Biophys. 1986 Jul; 248(1):305-23.
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• Wingrove DE, Amatruda JM, Gunter TE. Glucagon effects on the membrane potential and calcium uptake rate of rat liver mitochondria. J Biol Chem. 1984 Aug 10; 259(15):9390-4.
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• Gunter TE, Chace JH, Puskin JS, Gunter KK. Mechanism of sodium independent calcium efflux from rat liver mitochondria. Biochemistry. 1983 Dec 20; 22(26):6341-51.
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• Terman BI, Gunter TE. Characterization of the submandibular gland microsomal calcium transport system. Biochim Biophys Acta. 1983 Apr 21; 730(1):151-60.
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• Gunter KK, Gunter TE, Jarkowski A, Rosier RN. A method of resuspending small vesicles separated from suspension by protamine aggregation and centrifugation. Anal Biochem. 1982 Feb; 120(1):113-24.
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• Rosier RN, Tucker DA, Meerdink S, Jain I, Gunter TE. Ca2+ transport against its electrochemical gradient in cytochrome oxidase vesicles reconstituted with mitochondrial hydrophobic proteins. Arch Biochem Biophys. 1981 Sep; 210(2):549-64.
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• Puskin JS, Gunter TE, Coene MT. On the role of inorganic phosphate in divalent-cation sequestration by mitochondria. Eur J Biochem. 1980 May; 106(2):425-9.
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• Rosier RN, Gunter TE. Calcium uptake by cytochrome oxidase vesicles. FEBS Lett. 1980 Jan 1; 109(1):99-103.
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• Rosier RN, Gunter TE, Tucker DA, Gunter KK. A rapid method for separating small vesicles from suspension. Anal Biochem. 1979 Jul 15; 96(2):384-90.
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• Gunter TE, Rosier RN, Tucker DA, Gunter KK. Uptake of calcium and manganese by rat liver submitochondrial particles. Ann N Y Acad Sci. 1978 Apr 28; 307:246-7.
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• Gunter TE, Gunter KK, Puskin JS, Russell PR. Efflux of Ca2+ and Mn2+ from rat liver mitochondria. Biochemistry. 1978 Jan 24; 17(2):339-45.
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• Keith AD, Snipes W, Mehlhorn RJ, Gunter T. Factors restricting diffusion of water-soluble spin labels. Biophys J. 1977 Sep; 19(3):205-18.
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• Puskin JS, Gunter TE, Gunter KK, Russell PR. Evidence for more than one Ca2+ transport mechanism in mitochondria. Biochemistry. 1976 Aug 24; 15(17):3834-42.
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• Armbrecht HJ, Gunter TE, Puskin JS, Terepka AR. An electron paramagnetic resonance study of Mn2+ uptake by the chick chorioallantoic membrane. Biochim Biophys Acta. 1976 Mar 19; 426(3):557-69.
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• Armbrecht HJ, Terepka AR, Gunter TE. Energy-dependent Mn2+ and Ca2+ uptake by the embryonic chick chorioallantoic membrane. Biochim Biophys Acta. 1976 Mar 19; 426(3):547-56.
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• Gunter TE, Puskin JS. The use of electron paramagnetic resonance in studies of free and bound divalent cation: the measurement of membrane potentials in mitochondria. Ann N Y Acad Sci. 1975 Dec 30; 264:112-23.
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• Gunter RE, Puskin JS, Russell PR. Quantitative magnetic resonance studies of manganese uptake by mitochondria. Biophys J. 1975 Apr; 15(4):319-33.
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• Puskin JS, Gunter TE. Electron paramagnetic resonance of copper ion and manganese ion complexes with the ionophore A23187. Biochemistry. 1975 Jan 14; 14(1):187-91.
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• Puskin JS, Gunter TE. Ion and pH gradients across the transport membrane of mitochondria following Mn ++ uptake in the presence of acetate. Biochem Biophys Res Commun. 1973 Apr 2; 51(3):797-803.
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• Puskin JS, Gunter TE. Evidence for the transport of manganous ion against an activity gradient by mitochondria. Biochim Biophys Acta. 1972 Sep 20; 275(3):302-7.
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• Gunter TE, Puskin JS. Manganous ion as a spin label in studies of mitochondrial uptake of manganese. Biophys J. 1972 Jun; 12(6):625-35.
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• Gunter TE, Gunter KK. Pressure dependence of the helix-coil transition temperature for polynucleic acid helices. Biopolymers. 1972 Mar; 11(3):667-78.
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