Myoglobin translational diffusion in rat myocardium and its implication on intracellular oxygen transport.

A cornerstone of respiratory regulation stands on the capacity of myoglobin (Mb) to store O2 or to facilitate O2 transport. In marine mammals, the high concentration of Mb could certainly supply O2 during a dive or apnoea (Dolar et al. 1999; Guyton et al. 1995; Kooyman, 1998; Ponganis et al. 2002). In adaptation to high altitude, enhanced Mb expression increases the O2 depot (Gimenez et al. 1977; Terrados et al. 1990). These observations agree with the correlation between Mb concentration (O2 supply) and oxidative capacity in different species (Wittenberg & Wittenberg, 2003). Yet, in spontaneously beating rat heart, Mb can prolong normal heart function for only a few seconds (Chung & Jue, 1996). Without any Mb, neither myocardial nor skeletal muscle function suffers any apparent physiological impairment (Garry et al. 1998; Godecke et al. 1999). The physiology canon also states that Mb can facilitate O2 diffusion. In contrast to the low solubility of O2, the high O2 carrying capacity of Mb can confer an advantage in transporting O2 from the sarcolemma to the mitochondria (Wittenberg, 1970; Wittenberg & Wittenberg, 1989). In vitro studies have confirmed that O2 diffuses faster in solution containing Mb than in Mb-free solution. Mb exhibits sufficient mobility and O2 carrying capacity to compete effectively with free O2 (Johnson et al. 1996). In vivo, however, the contribution of Mb-facilitated O2 diffusion remains unclear. Without a definitive translational diffusion coefficient for Mb (DMb) in vivo, the theory of Mb-facilitated diffusion languishes for experimental confirmation. Over the years, researchers have attempted to estimate endogenous Mb diffusion in the cell by measuring Mb diffusion in concentrated solution, in tissue homogenate and in myoglobin-free frog muscle (Moll, 1968; Riveros-Moreno & Wittenberg, 1972; Baylor & Pape, 1988). The results have varied widely. Fluorescence recovery after photobleaching (FRAP) techniques have recently tracked the photoxidation of Mb in superfused rat diaphragm or the diffusion of microinjected modified Mb in isolated muscle fibre. These experiments have determined a low DMb that cannot support any significant role for Mb in facilitating O2 diffusion (Jurgens et al. 1993; Papadopoulos et al. 2001). However, the FRAP experiments do not actually measure endogenous Mb diffusion and utilize model systems that do not adequately mimic respiring tissue (Groebe, 1995). Moreover, they disagree with the in vivo NMR observation of Mb rotational diffusion, which predicts a much faster DMb (Livingston et al. 1983; Wang et al. 1997). Because the 1H NMR can detect the distinct γCH3 Val E11 signal of MbO2 in myocardium at −2.8 ppm, an opportunity exists to apply pulsed-field gradient technique to map endogenous Mb translational diffusion in perfused myocardium (Stejskal & Tanner, 1965; Kreutzer et al. 1992). Indeed, MbO2 diffuses with an average coefficient about 4 times faster than the FRAP-determined diffusion coefficient and shows no orientation preference. The DMb also matches precisely the value predicted by the NMR rotational diffusion analysis (Wang et al. 1997). Given the Mb concentration in tissue, the partial pressure of O2 that half saturates Mb (P50) and the DMb, the analysis establishes an equipoise diffusion PO2 in the cell, where Mb and free O2 contribute equally to O2 transport. In the basal state rodent myocardium or skeletal muscle, Mb cannot play a significant role in facilitating O2 diffusion. In contrast, marine mammal muscle with a high Mb concentration can utilize Mb-facilitated O2 diffusion under all physiological conditions. The conclusion agrees with Mb studies in which rat myocardium inhibited with a significant fraction of CO-bound Mb exhibits no sign of respiration or metabolism impairment (Glabe et al. 1998; Chung et al. 2006). The NMR-determined translational diffusion coefficient of endogenous Mb in respiring myocardium has established a key parameter that does not lend support to the hypothesis that Mb has an overall general role in facilitating O2 transport in respiring tissue. Instead, the DMb defines the physiological conditions in which Mb can contribute significantly to the intracellular O2 flux.