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Comparative Study
. 2006 Aug 15;575(Pt 1):191-200.
doi: 10.1113/jphysiol.2006.114116. Epub 2006 Jun 1.

Altered energy transfer from mitochondria to sarcoplasmic reticulum after cytoarchitectural perturbations in mice hearts

James R Wilding  1 , Frédéric Joubert, Carla de Araujo, Dominique Fortin, Marta Novotova, Vladimir Veksler, Renée Ventura-Clapier
Affiliations
Comparative Study

Altered energy transfer from mitochondria to sarcoplasmic reticulum after cytoarchitectural perturbations in mice hearts

James R Wilding et al. J Physiol. .

Abstract

Sarcoplasmic reticulum (SR) calcium pump function requires a high local ATP/ADP ratio, which can be maintained by direct nucleotide channelling from mitochondria, and by SR-bound creatine kinase (CK)-catalysed phosphate-transfer from phosphocreatine. We hypothesized that SR calcium uptake supported by mitochondrial direct nucleotide channelling, but not bound CK, depends on the juxtaposition of these organelles. To test this, we studied a well-described model of cytoarchitectural disorganization, the muscle LIM protein (MLP)-null mouse heart. Subcellular organization was characterized using electron microscopy, and mitochondrial, SR and myofibrillar function were assessed in saponin-permeabilized fibres by measuring respiration rates and caffeine-induced tension transients. MLP-null hearts had fewer, less-tightly packed intermyofibrillar mitochondria, and more subsarcolemmal mitochondria. The apparent mitochondrial Km for ADP was significantly lower in the MLP-null heart than in control (175 +/- 15 and 270 +/- 33 microM, respectively), indicating greater ADP accessibility, although maximal respiration rate, mitochondrial content and total CK activity were unaltered. Active tension in the myofibres of MLP-null mice was 54% lower than in controls (39 +/- 3 and 18 +/- 1 mN mm(-2), respectively), consistent with cytoarchitectural disorganization. SR calcium loading in the myofibres of MLP-null mice was similar to that in control myofibres when energy support was provided via Bound CK, but approximately 36% lower than controls when energy support was provided by mitochondrial (P < 0.05). Mitochondrial support for SR calcium uptake was also specifically decreased in the desmin-null heart, which is another model of cytoarchitectural perturbation. Thus, despite normal oxidative capacity, direct nucleotide channelling to the SR was impaired in MLP deficiency, concomitant with looser mitochondrial packing and increased nucleotide accessibility to this organelle. Changes in cytoarchitecture may therefore impair subcellular energy transfer and contribute to energetic and contractile dysfunction.

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Figures

Figure 1
Figure 1. Electron microscopic images of left ventricle from control and MLP-null mice
A, overview of a control myocyte in longitudinal section, with mitochondria arranged in longitudinal columns. B, overview of an MLP-null cardiomyocyte, showing myofibrillar disorganization, an irregular arrangement of intermyofibrillar mitochondria, and increased content of subsarcolemmal mitochondria. C, detail of sarcomeres in a control myocyte, showing mitochondria tightly packed with SR. D, detail of sarcomeres in an MLP-null myocyte, showing looser packing of mitochondria with SR, irregular and widened Z-lines, and cytoplasm in the intermyofibrillar space. Arrowheads show mitochondria, arrows show SR.
Figure 2
Figure 2. Electron microscopic images of intercalated disks in control and MLP-null mouse heart, and of mitochondrial aggregates in MLP-null myocytes
A, longitudinal section of control myocyte showing myofibrils attached to the intercalated disc. B, an oblique section through the widely spread intercalated disc of an MLP-null myocyte. C, a longitudinal section through a region of intercalated disc lacking myofibrils in an MLP-null myocyte. D, mitochondrial aggregates in MLP-null myocytes.
Figure 3
Figure 3. Representative tension transients during SR calcium release in control (A) and MLP-null (B) cardiac fibres
The SR was loaded with calcium for 3 min in permeabilized fibres under different energetic conditions (ATP + Mito + CK, ATP + Mito, ATP + CK, ATP, see Methods), and then calcium load was estimated by relating these tension transients, elicited by caffeine stimulation, to the calcium–tension curve for each fibre.
Figure 4
Figure 4. SR calcium loading over time under different energetic conditions in control and MLP-null mouse cardiac fibres
SR calcium load was estimated using the surface of the tension transient caused by caffeine stimulation (SCa). Energy supplied by exogenous ATP, mitochondria and the creatine kinase reaction (ATP + CK + Mitro, panel A), by exogenous ATP and mitochondria (ATP + Mitro, panel B), by exogenous ATP and creatine kinase (ATP + CK, panel C) and by exogenous ATP alone (ATP, panel D). *P < 0.05 versus control (Con).
Figure 5
Figure 5. Relative SR calcium content after loading for 180 s, as a proportion of the load effected by ATP + Mito + CK in the same fibre, under different energetic conditions in control, MLP-null and desmin-null cardiac fibres
*P < 0.05 versus ATP + Mito + CK, †P < 0.05 versus ATP + CK, ‡P < 0.05 versus ATP + Mito. See Results for precise P values.
See this image and copyright information in PMC

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