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This report details histopathological findings of atypical vacuolated hepatocytes in the white perch Morone americana. Vacuolated cell lesions have been reported from a number of species in association with degraded habitats, often in the presence of hepatic neoplasms. Extensive atypical vacuolated cell lesions assuming characteristic acinar or ductular configurations have been reported previously only from the winter flounder Pseudopleuronectes americanus. Light microscopic findings, utilizing a variety of histochemical stains, suggest the identical nature of this lesion to that of winter flounder. The discovery of this lesion in a free-swimming species, feeding throughout the water colun~n, is in direct contrast to that of the bottom dwelling pleuronectid, feeding from and constantly in contact with potentially contaminated sediments. While the winter flounder has become the sentinel fish species for pollution monitoring on the northeast coast, increased surveillance of other species may prove atypical vacuolated cell lesions represent a common mechanism of chronic cellular injury in relation to impacted environments. Vacuolated cell lesions, hepatic and biliary, have been reported from a number of species under natural and experimental conditions (Smith et al. 1979, Hendricks et al. 1984, Cormier 1986, Grizzle & Thiyagarajah 1988, Hinton et al. 1988). In Boston Harbor (USA) a unique lesion was reported in winter flounder Pseudopleuronectes americanus by Murchelano & Wolke (1985); extensive areas of hepatocytes appeared replaced by atypical vacuolated cells in distinct acinar and ductular configurations. Unlike other reported vacuolated hepatocyte lesions, hepatocellular steatosis and clear cell foci, affected hepatocytes of winter flounder contain a condensed apical nucleus and a single large clear vacuole devoid of lipid, glycogen or mucopolysaccharides. The lesion also lacks the flocculent, eosinophilic material generally associated with cystic parenchyma1 degeneration or the locular appearance of spongiosis hepatis. Additionally, while extensive areas of hepatocytes may be replaced by hypertrophic vacuolated cells, often surrounded by connecO Inter-Research/Printed in Germany tive tissue, the lesion is restricted to the hepatic cords and normal architecture is generally preserved. Since 1985, several authors have reported this lesion in winter flounder from various sites along the New England seaboard, typically in association with contaminated sediments adjacent to population centers (Zdanowicz et al. 1986, Gardner et al. 1989). Similar lesions have been noted in rock sole Lepidopsetta bilineata, starry flounder Platichthys stellatus and white croaker Genyonemus lineatus on the west coast (M. S. Meyers, National Marine Fisheries Service, Seattle, WA, pers. comm.). Recently, a n atypical vacuolated hepatocyte lesion resembling that of winter flounder was found in one of a group of 5 white perch collected from a Rhode Island coastal salt pond open indirectly to Narragansett Bay. White perch, unlike winter flounder, are found in these shallow ponds throughout the year. While capable of tolerating full seawater, they prefer the brackish waters of the ponds, where they wander and feed in small schools. In winter they congregate in deeper waters to hibernate, running up into fresh or slightly brackish water to spawn April through June. White perch are highly predaceous, feeding on the spawn and fry of numerous fish and crustaceans (Bigelow & Schroeder 1953). Materials and methods. Five post-spawning female white perch Morone americana were taken by fyke net in July of 1990 in a population survey of coastal salt ponds of Rhode Island, USA. Fish were euthanized in tricaine methanesulfonate (MS-222), weighed, measured and necropsied immediately. Collected tissues were identified, fixed in 10 % neutral buffered formalin, embedded in paraffin and sectioned at 6 pm for light microscopic examination. Staining procedures, a s outlined by Luna (1968), included hematoxylin and esoin (H & E), periodic acid Schiff (PAS), Perls' method 226 Dls. aquat. Org. 11: 225-228, 1941 for iron, >lasson's trichrome (hlT), Ziehl-Neelsen acid fast stain (ZN) , and alcian blue (AB). Formalin fixed tissue was embedded in gelatin, seclioned frozen and stained for lipids with oil red 0 (ORO) and sudan black B (SB). Results and discussion. Fish appeared to be in good flesh, ranging in weight from 154 to 506 g and in length from 26 to 32 cm. Little abdominal fat was present. but was not considered unusual for post-spawning individuals. Livers were of firm consistency and uniform light mahogany coloration. Gross observation revealed the presence of multiple 1 mm white foci scattered throughout the hepatic and splenic parenchymas of all fish examined. Microscopically, these were identified as the metacercariae of digenetic trematodes. Metacercariae were encircled by a thln fibrous capsule, but did not appear to elicit a significant inflammatory response. Under light microscopy, prespawning individuals r e v ~ a l arge polygona! hepatocytes arrazged in irrzgular cords. The pattern is typical tubulosinusoidal, lacking distinct lobulation, as described in rainbow trout by Hampton e t al. (1985). Nuclei are spherical and generally centrally located with dispersed chromatin and single prominent nu.cleoli. Most hepatocytes contain one to several inclusions within their pale, finely granular cytoplasm. Small inclusions are (PAS) positive for glycogen, while larger inclusions, often perinuclear, are (SB) and (ORO) positil-C: for lipids. In contrast, hepatocytes of post-spawning indi\.iduals appeared small, darker staining and dcpleted of cytoplasmic inclusions. Shrunken hepatocytes result In an appearance of dilated sinusoids. Atypical vacuolated hepatocytes, possessing a single large clear vacuole, occurred singly or in characteristic acinar (Fig. 1) or ductular (Fig. 2) patterns. No histological alterations were noted in hepatocytes adjacent to the vacuolated foci, nor were there any associated inflammatory changes. Nuclei were compact, uniformly basophilic and displaced apically. Occasionally, small PAS positive inclusions were present, but invariably displaced peripherally by the large central vacuole. The extensive areas of vacuolated cells were often invested in a fine sheath of connective tissue (MT). Vacuoles were consistently negative for glycogen (PAS), mucopolysaccharides (AB) and lipids (SB, CRC) as in winter flounder. In addition to the vacuolated cell lesion, histopathological changes in the affected liver included multiple foci of necrosis, focal hepatitis, pericholangitis and vasculitis. Vacuolated cells appeared random in distribution, demonstrating no spatial relationships to any of the aforementioned lesions or metacercanae. Pericholangi.al fibrosis was common. A single focus of bile duct hyperplasia also contained vacuolated cells Fig. 1. Morone americana. Acinus of vacuolated hepatocytes demonstrating large c ~ n t r a l vacuoles and apicslly displaced nuclei. Note investing sheath of collagcnous fibers. H & E; X 1000 Camus S \Volkc ..\typical ht-patlc crll l e s~on in white perch 227 Fig. 2. Morone amedcana. Vacuolated cells occurring singly and in a large ductular configurat~on H & E; X 500 within its connective tissue matrix, light microscopically indistinguishable from those found in the hepatic parenchyma. Pigmented macrophage aggregates were numerous, strongly (ZN) and (PAS) positive, but only weakly Pei-l's positive for ferric iron. Irregular focl of coagulat~ve necrosis contained amorphous hyalin material and s~ng le pigment laden macrophages with staining properties identical to those of nearby aggregates. Tissue coarse eosinophilic granulocytes were also common. Number, extent and severity of pathological lesions appeared to increase dramatically with fish size, even within this small sample group. The vacuolated cell lesion was present in the second largest fish, a 480 g specimen. The ultrastructural characteristics of this lesion in winter flounder have recently been reviewed (Bodammer & Murchelano 1990). The large vacuole has been determined to represent the dilated cisternae of a modified endoplasmic reticulum system. These cells show additional signs of chronic cellular injury, including numerous autophagic vacuoles and condensed mitochondna. At present, the cellular origin of this lesion and its progression remains unclear Based on cellular orientation and junctional complexes, it appears to involve both bile preductular cells (Moore et al. 1990) and hepatocytes (Bodammer & Murchelano 1990). Likewise, the relationship of this lesion to hepatic neoplasia is also uncertain. Flounder possessing atypical vacuolated cell lesions demonstrate a high incidence of hepatic neoplasia (Murchelano & Wolke 1985, 1991). Statistical evidence supports a correlation between hepatic neoplasia in flatfish and polynuclear aromatic hydrocarbons (Malins et al. 1984, Gardner et al. 1989). While serial sections of the vacuolated cell llver did not demonstrate neoplasms, it did possess a variety of lesions associated with polluted environments (Meyers et al. 1987, Murchelano & Wolke 1991). Demonstration of an atypical vacuolated hepatocyte lesion in the white perch suggests a common mechanism of cellular injury, not unique to the winter flounder or solely bottom dwelling species. While experimental evidence is lacking, the striking cellular alterations are presumed to be the result of environmental toxins. The nature of the vacuolar contents in both species is unclear, but histochemical staining has shown them to be distinct from the more well described lipid and polysaccharide inclusions seen in other fish. Electron microscopy, in winter flounder, has identified the vacuoles as dilated cisternae of endoplasmic reticulum, often containing membranous inclusions considered autophagic in nature, a significant sign of sublethal cell injury (Bo

Atypical hepatic vacuolated cell lesion in the white perch Morone Americana