Proceedings of The Physiological Society

Europhysiology 2018 (London, UK) (2018) Proc Physiol Soc 41, PCB226

Poster Communications

Effects of ouabain and/or salbutamol on subcellular glycogen distribution in isolated rat soleus muscle

J. Nielsen1, N. Ørtenblad1

1. Department of Sports Science and Clinical Biomechanics, University of Southern Denmark, Odense M, Denmark.

  • Figure 1. Glycogen for the Na+,K+ pumps: Where does it come from?Transmission electron microscopy reveals the subcellular localization of glycogen (black dots). The Na+,K+ pumps of the t-system (yellow arrow) are located distant both to glycogen particles located within the myofibrils (Intra glycogen, blue circle) and to those located close to sarcoplasmic reticulum (IMF glycogen, red circle). Scale bar = 100 nm.

In skeletal muscle, glycogen is essential for sustaining prolonged work, however the underlying mechanism(s) remains unknown (Allen et al. 2008; Ørtenblad et al. 2013). One hypothesis is that compartmentalized energy transfer is required from the breakdown of specific glycogen particles located in a juxtaposition to specific energy-dependent functions creating a spatially separate and efficient energy transfer network (Dzeja et al. 2007; Nielsen et al. 2009). Here, we aimed to investigate how salbutamol (a β2-agonist, which stimulates glycogenolysis) and ouabain (a cardiac glycoside, which inhibits Na+,K+ pumps) influenced spatially distinct depots of glycogen particles in skeletal muscle fibres. Soleus muscles from male sprague dawley rats (n = 26, 164 ± 17 g (mean ± SD)) were incubated in a Krebs-Ringer solution with 5 mM glucose (pH 7.4) at 25 oC, and continuously gassed with 95% O2 and 5% CO2. Paired muscles (from left and right leg) were randomly allocated to two of the four groups: control (C; n = 11), salbutamol (S; 10 µM, n = 15), ouabain (O; 1 mM; n = 11) or ouabain + salbutamol (OS; n = 15). Transmission electron microscopy was conducted to quantitate the volumetric content of glycogen particles in three distinct locations (Figure 1): 1) intermyofibrillar (IMF, 70% of total), 2) intramyofibrillar (Intra, 25% of total), and 3) subsarcolemmal (SS, 5% of total). Data are presented as medians and interquartile range. Main and interaction effects were tested using a linear mixed-effects model. In IMF glycogen, salbutamol mediated a 40% decrease (S: 3.44 (2.77:4.62) vs C: 5.76 (4.82:6.91), which was not affected (a 42% decrease) by pre-treatment with ouabain (OS: 4.48 (3.30:6.14) vs O: 7.71 (4.62:9.13), main effect: P < 0.0001, interaction: P = 0.24). Ouabain-treatment alone increased IMF glycogen by 10% (OS+O: 5.01 (3.28:7.83) vs C+S: 4.54 (3.15:6.02), P = 0.009). In Intra glycogen, salbutamol mediated a 55% decrease (S: 1.45 (0.71:2.34) vs C: 3.21 (2.06:4.77), which was slightly attenuated (to a 48% decrease) by pre-treatment with ouabain (OS: 1.92 (1.49:2.52) vs O: 3.67 (2.00:5.18), main effect: P = 0.0001, interaction: P = 0.096). Ouabain-treatment alone did not significantly affect Intra glycogen (OS+O: 2.39 (1.50:3.70) vs C+S: 2.06 (1.09:3.21), P = 0.23). Neither salbutamol (P = 0.31), ouabain (P = 0.97) nor both combined (P = 0.65) influenced SS glycogen (S: 3.60 (2.03:5.85) vs C: 4.47 (2.74:7.21) and (OS: 4.31 (2.79:6.17) vs O: 4.81 (2.86:6.59). In conclusion, stimulation of glycogenolysis by salbutamol mediates a compartmentalized utilization of glycogen depots as characterized by a decrease in both IMF and Intra glycogen, but not SS glycogen. Blocking Na+,K+ pumps with ouabain, tended to attenuate the utilization of Intra glycogen. The present data suggest that Na,K-pumps preferentially use this small depot of Intra glycogen, located in the sarcomeric I-band near the t-system.

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