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Background and Objective: To gain some insight into the photostimulation of isolated hepatocytes irradiated with Helium-Neon (He-Ne) laser light certain biochemical events were studied with respect to two mechanisms: i) the direct light dependent activation of certain biochemical events investigated in intact cells and isolated mitochondria, ii) the indirect stimulation of processes per se light independent., Study Designs/materials and Methods: Irradiation of either isolated hepatocytes or isolated rat liver mitochondria was carried out with He-Ne laser (wavelength, 632.8 nm; fluence, 0.24 J cm-2; fluence rate, 12 mW cm-2). Changes in mitochondrial membrane potential in isolated hepatocytes were monitored using the cationic probe safranine. The c-fos expression was studied by Northern blot and immunoblot analysis., Results: As a result of irradiation, increase of the mitochondrial membrane potential was found to occur in irradiated hepatocytes both in the presence or in the absence of CaCl2. The hyperpolarization of the mitochondrial membrane is assumed to cause an increase in mitochondrial Ca2+ uptake that was measured in isolated mitochondria. Finally, an increase in c-fos expression was found in irradiated hepatocytes when incubated in the presence of CaCl2., Conclusion: This paper gives additional information on the mechanism by which He-Ne laser light, either directly or in a cascade-like effect dependent on increase in cell Ca2+, can cause cell stimulation.

Isolated hepatocytes were irradiated with Helium-Neon laser (fluence: 0.24 Joules x cm-2, fluence rate: 12 mW x cm-2) and changes of both cytosolic free Ca2+ concentration and cell membrane potential were checked by measuring fura-2 and bis-oxonol fluorescence respectively. Irradiation resulted in an enhancement in cytosolic free Ca2+ concentration that requires the presence of Ca2+ in the phase outside hepatocytes; consistently an increase in cell membrane potential was measured correlated with it. Interestingly, the rate of increase of both cytosolic free Ca2+ concentration and cell membrane potential shows special time dependent features similar to those peculiar of oscillatory processes.

In order to gain an insight into the mechanism of cell photostimulation by laser light, protein synthesis was measured in hepatocytes irradiated with a low-power, continuous-wave He-Ne laser (fluence, 0.24 J cm(-2); fluence rate, 7 and 12 mW cm(-2)). As a result of irradiation, the rate and amount of 35S-methionine incorporated into newly synthesized proteins increased, as demonstrated by gel electrophoresis and quantitative analysis of labelled protein bands. The stimulation of protein synthesis was fluence dependent, with a maximum stimulation at 0.24 J cm(-2) for both fluence rates (12 and 7 mW cm(-2)). Both cytosolic and mitochondrial protein synthesis increased as a result of irradiation, as demonstrated by the measurement of hepatocytes previously treated with chloramphenicol and cycloheximide respectively. An initial investigation showed that stimulation of protein synthesis also occurred in hepatocytes irradiated with a non-coherent radiation source (fluence, 0.24 J cm(-2)).

In order to gain a degree of insight into the mitochondrial component/s responsible of the mitochondria-red light interaction, isolated rat liver mitochondria were irradiated with a Helium-Neon laser (energy dose 2 Joules/cm2, light power 10 mW) and measurements made of the activity of cytochrome c oxidase. A low, but statistically significant increase in the oxygen uptake was found, as polarographically measured, in the presence of rotenone and antimycin A, with ascorbate and TMPD used as substrate pair. Measurements were also made both of the electron transfer and of proton pumping activity: as a result of a major stimulation in the proton pumping activity, about 55% increase of <--H+/e- ratio was found in irradiated mitochondria.