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We developed an easy to use and non-invasive method to study sarcomere motion of enzymatically isolated myocytes which can be simultaneously combined with auxotonic force detection, thus being very useful when studying the contractile performance of cardiac cells. This method basically consists in analyzing the periodicity of the cell striation pattern using the Cooley-Tukey fast Fourier transform (FFT) algorithm on a video image of the cell during the course of the experiment. A longitudinal fraction of the cell image is recorded with a CCD TV camera, digitized, then transiently stored on a computer and used to calculate the spectrum corresponding to the distribution of the sarcomere lengths (SL). The method gives a real-time measurement of the most probable value of sarcomere length in one isolated cell with a temporal resolution of 20 ms. When used on a cell attached between two carbon fibers, the auxotonic force developed by the cell upon electrical stimulation can be simultaneously measured together with the SL in various conditions of stretch. Preliminary results have been presented in abstract form (Gannier et al., vol 24, pp. S47, 1992).

Objectives: The aim of the study was to investigate the mechanisms responsible for provoking and maintaining a large, stretch-induced, increase in the level of resting calcium in single guinea-pig ventricular myocytes. In particular, we wished to test the relative importance of intracellular and extracellular sources of calcium in this phenomenon. Methods: Carbon fibres were used to stretch cells loaded with the fluorescent calcium indicator Indo-1. Sarcomere length and internal calcium activity ([Ca2+]i) were measured. Experimental results from our present and previous studies were compared with those predicted by the OXSOFT HEART (version 4) model of the guinea-pig ventricular myocyte incorporating a stretch-activated channel. Results: The stretch-induced increase in [Ca2+]i was found to be sensitive to removal of [Ca2+]0 and application of the Ca2+-channel blocker verapamil (1 μM). The phenomenon was not sensitive to disruption of sarcoplasmic reticulum function by ryanodine (1 μM) nor to the Na+ channel blocker TTX (30 μM). Our experimental findings were reproduced in the modelling study. Conclusions: The stretch-induced increase in [Ca2+]i is modulated by extracellular sources of Ca2+ rather than intracellular Ca2+ stores and is not indiscriminately sensitive to blockers of depolarizing current. We propose that the stretch-induced increase in [Ca2+]i may be triggered by activation of stretch-activated channels but that a combination of stretch-activated current and Ca2+-window current maintain the increased levels of resting [Ca2+]i. [ABSTRACT FROM PUBLISHER]

The Frank-Starling mechanism is one of the most fundamental properties of cardiac muscle for exerting intrinsic control over contractile force on a beat to beat basis. According to this law, the systolic contractile force depends on the preceding diastolic blood loading. In other words stretching the ventricular muscle induces an increase in contractility. In multicellular preparations the mechanisms that underlie the length-force relationship in the myocardium have been the subject of considerable discussion and controversy.1-7 [ABSTRACT FROM PUBLISHER]

Objective: The aim was to test the hypothesis that in single guinea pig ventricular myocytes a large stretch induced increase in resting calcium was sensitive to the mechanosensitive channel blocker streptomycin. Methods: Carbon fibres were used to stretch cells loaded with the fluorescent calcium indicator indo-1. Force, sarcomere length, and internal calcium activity ([Ca2+]i) were measured. Results: In approximately 60% of the cells studied, a stretch which increased sarcomere length by approximately 6% caused a large increase in [Ca2+]i (up to 60% of the size of a [Ca2+]i transient at 0.25 Hz). When a mixture of antibiotics (streptomycin-penicillin) was used in solutions to isolate and store cells, this phenomenon was never observed (n = 19 cells). Direct application of physiological saline solution (PSS) could not reverse the increase in [Ca2+]i within 60 s of application (n = 7 cells). Direct application of penicillin [1000 IU per 50 ml (40 μM)] reversed the increase in [Ca2+]i within 60 s of application in only 3/7 cells. In contrast direct application of the aminoglycoside antibiotic streptomycin (40 μM) rapidly reversed the large increase in [Ca2+]i induced by stretch in each of 13 cells [within 18(SD 10) s of application]. Acute application of 40 μM streptomycin did not modify L-type Ca2+ currents measured under whole cell patch clamp conditions. Measurement of the resting tension – sarcomere length curves in cells stored in solution containing streptomycin and penicillin revealed two populations of cells on the basis of their stiffness. Conclusions: This stretch induced increase in [Ca2+]i may be associated with stretch activated arrhythmias in the heart. The effects of streptomycin are consistent with its reported inhibitory action on stretch activated channels.Cardiovascular Research 1994;28:1193-1198 [ABSTRACT FROM PUBLISHER]