The hepatocytes of the brown trout (Salmo trutta f. fario): a stereological study of their number and size during the breeding cycle

A firm knowledge of the normal structure is crucial for evaluating pathological processes and morphofunctional correlations. Stereological liver structure characterization had its debut for mammals in the 1960s, but only in the 1980s did it start to be used in fishes. Using stereology, our aim was to verify the hypothesis that in parallel with the well-known annual seasonal changes in the liver-body ratio of brown trout, hepatocytes would vary their number and/or size, and that gender differences likely exist. Three-year-old specimens were used. Five animals per gender were examined in May (endogenous vitellogenesis), September (exogenous vitellogenesis), and February (spawning season end). The liver was fixed by perfusion, and its total volume estimated. Systematically sampled material was embedded in epoxy or in metachrylate resins. Stereology was executed on light and electron microscopy images. Unbiased design-based techniques were applied, using physical disectors and differential point counting. Target parameters were the relative (per unit volume) and total number of hepatocytes, the mean cell and nuclear volumes, and the total volumes of hepatocytes and their nuclei. Data support that in both genders the number of hepatocytes and the volume of its nucleus change along the breeding cycle. The cell number increased from endogenous to exogenous vitellogenesis (accompanying relative liver size gains), later followed by a decline in the cell number, still detectable after the spawning season. The total liver volumes of the cell and nucleus also increased from May to September in females, despite that the mean hepatocyte nuclear volume showed a minimum in September. No statistical changes in the mean cell volume were detected, regardless of the tendency for lower mean values in September. Changes were more marked in females and showed a higher correlation with the gonad weight. It was firstly suggested that numerical (rather than cell size) changes govern the shifts of the relative liver weight seen during the brown trout annual breeding cycle, and eventually of other fishes. We hypothesized that there are seasonal cycles of hepatocyte mitosis (from after spawning to exogenous vitellogenesis) and of apoptosis (at spawning). These cycles would be regulated by sex steroids, being more striking in females.