An in vivo model for studying the dynamics of intracellular free calcium changes in slow- and fast-twitch muscle fibres.
The understanding of the regulation of the free cytosolic [Ca2+] ([Ca2+]i) in skeletal muscle is hampered by the lack of techniques for quantifying free [Ca2+]i in muscle fibres in situ. We describe a model for studying the dynamics of free [Ca2+]i in the fast-twitch extensor digitorum longus (EDL) and the slow-twitch soleus (SOL) muscles of the rat in vivo using caffeine superfusion to induce changes in free [Ca2+]i. We assumed that differences in sensitivity between the two muscle types for this substance reflect differences in intracellular Ca2+ handling in the fibres of which these muscles consist. The Indo-1 ratiometric method, using intravital microscopy with incident light, was adapted to measure free [Ca2+]i in vivo. Fluorescence images were collected by means of a digital camera. Caffeine superfusion at 37 degrees C for 2 min, at concentrations of 1, 2, 5, 10 or 20 mmol/l, induced a concentration-dependent increase in free [Ca2+]i and revealed differences in caffeine sensitivity between the muscle types, with the SOL being more sensitive. In a separate set of experiments the contracture threshold, as assessed by topical application of caffeine, was determined in both muscle types. EDL had a higher threshold for developing contracture than SOL. These finding are in agreement with previous in vitro studies. We may conclude that the dynamics of free [Ca2+]i can be assessed reliably in intact mammalian muscle in vivo.
Bátkai
S
S
2nd Department of Physiology, Semmelweis Medical University, Budapest, Hungary. batkai@elet2.sote.hu
Rácz
I B
IB
Ivanics
T
T
Tóth
A
A
Hamar
J
J
Slaaf
D W
DW
Reneman
R S
RS
Ligeti
L
L
D000818:Animals; D002110:Caffeine / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D005456:Fluorescent Dyes; D007211:Indoles; D008297:Male; D008954:Models, Biological; D009119:Muscle Contraction / Q000187:drug effects; D018656:Muscle Fibers, Fast-Twitch / Q000187:drug effects / Q000378:metabolism*; D018657:Muscle Fibers, Slow-Twitch / Q000187:drug effects / Q000378:metabolism*; D018482:Muscle, Skeletal / Q000378:metabolism* / Q000648:ultrastructure; D051381:Rats; D017207:Rats, Sprague-Dawley
D005456:Fluorescent Dyes; D007211:Indoles; D002110:Caffeine; C048960:indo-1; D002118:Calcium