Vol 159, No 1 (2021)

AIM: The work aimed to reveal structural signs of autophagy in the cytoplasm of isolated hepatocytes in the dynamics of their cultivation.

MATERIALS AND METHODS: The cultivated hepatocyte culture cell cycle was studied by flow cytofluorometry. The cells were cultured for 1, 24, and 48 hours. Morphometric analysis was performed using of the computer program Image J. The diameters of the nuclei and cytoplasm of hepatocytes, the volumes of nuclei and cytoplasm, and the nuclear-cytoplasmic ratio were determined. The concentration of intracellular organelles and autophagy was evaluated with magnification by 30000 times.

RESULTS: The cell cycle arrest in the G₀/G₁ stage after 24 hours of hepatocyte cultivation and the preservation of their viability by hour 48 of the experiment without increase in the percentage of cells in the apoptosis stage were revealed. The decrease in the absolute count of cells was registered, as well as an increase in the nuclear-cytoplasmic ratio indicating a decrease in the proportion of hepatocyte cytoplasm in the course of cultivation. After 24 hours of cultivation, autophagosomes with fragments of cytoplasm, glycogen rosettes, and autolysosomes with partially degraded material were revealed in the cell cytoplasm. By hour 48 of the study, a significant decrease in the volume density of glycogen and mitochondria was noted, as well as an increase in basal autophagy in hepatocytes, with a prevalence of glycophagy and mitophagy.

CONCLUSIONS: Autophagy maintains cellular homeostasis of isolated hepatocytes under standard culture conditions, as evidenced by a decrease in the volume density of glycogen and mitochondria, and an increase in basal autophagy in the hepatocyte cytoplasm. The findings indicate the contribution of autophagy to the survival of the primary culture of hepatocytes and can be used as an indicator of the adequacy of culturing conditions.

AIM: The work aimed to investigate inducible NO synthase (iNOS) expression in dentate gyrus in mature rats when modeling depression, as well as the establish the pharmacological correction possibility of detected changes with Phenibut and compounds under laboratory codes of RSPU-189, RSPU-135.

MATERIALS AND METHODS: Depressive-like behavior in animals was modeled by combining stressful stimuli such as loud sound, pulsating bright light, and vibration simultaneous with constant restriction of mobility and fluctuations in temperature of environment for 7 days (daily for 30 minutes). Changes in level of iNOS expression in dentate gyrus were assessed by calculating relative area of immunoreactive material (IRM) and staining intensity in points from 0 to 3.

RESULTS: Compared with the control group, rats with experimental depression showed an increase in expression of iNOS-IRM in cytoplasm of neuronal perikarya in granular layer of dentate gyrus, as well as an increase in relative area of iNOS-IRM in neuropil and nerve cells. The use of the compound RSPU-189 (salifen) demonstrated to a greater extent the corrective effect, since in the cytoplasm of neuronal perikarya in granular layer of dentate gyrus of rats, there was a decrease in the expression of iNOS-IRM, as well as a decrease in the relative area of iNOS-IRM in neuropil and nerve cells, which corresponded to values of these parameters in the control group of animals.

CONCLUSIONS: An experimental modeling of depression in dentate gyrus of mature rats revealed an increase of iNOS-IRM expression, the decrease of which was noted in its pharmacological correction with the compound RSPU-189 (salifen), which may indicate the predominant neuroprotective effect of this compound on GABAergic neurotransmission mechanisms.