352 



Fishery Bulletin 104(3) 



D E F 



Figure 2 



Photomicrographs of histological cross-sections of rex sole iGIyptocephalus zachirus) ovaries: (A) early perinuclear 

 (EP) oocytes and late perinuclear [EP] oocytes, with a large central nucleus <N), that have not begun vitellogenesis; 

 (B) early yolked oocyte beginning vitellogenesis with yolk globules lYG) at periphery of oocyte; (C) advanced 

 yolked oocyte; (D) migratory-nucleus oocyte; (E) hydrated oocytes with fused yolk (FYl; (F) postovulatory follicles 

 (POPs). Each photomicrograph has a scale bar equal to 0.1 mm. 



were analyzed for age determination. The left otolith, 

 or blind-side otolith, was read because it is thicker 

 along the proximal-distal axis, thereby allowing the 

 annuli to be less compressed and easier to differenti- 

 ate when counted from the core to the otolith edge 

 along the sulcus (Anderl^). Otoliths were processed 

 with standard break-and-burn techniques (Beamish 

 and Chilton, 1982). Two experienced readers each read 

 117 otoliths and had initial agreement of 67%. The 

 magnitude of the initial discrepancies within an age- 

 group ranged from -2 to 4-3 years for females from age 

 2 to 18 years. All discrepancies of age estimates were 

 resolved between readers through combined analysis. 

 Once all discrepancies were resolved, the remaining 

 otoliths were analyzed. 



Histological analysis 



Tissue from the middle portion of the right (ocular-side) 

 ovary was embedded in paraffin Paraplast (McCormick 

 Scientific, St. Louis, Missouri). Serial sections (5-8 



^ Anderl, D. 2003. Personal commun. Alaska Fisheries 

 Science Center, National Marine Fisheries Service, 7600 

 Sand Point Way NE, Seattle, WA 98115. 



microns thick) were prepared and stained with Harris 

 hematoxylin (Sigma Aldrich, St. Louis, MO) followed 

 by eosin counterstain (Sigma Aldrich, St. Louis, MO) 

 (H&E). Each ovary section was examined for the pres- 

 ence or absence of the following oocyte stages (see Fig. 2): 

 oocytes that had not begun vitellogenesis (early perinu- 

 clear [EP] and late perinuclear [LP]); oocytes in the first 

 vitellogenic stages (early yolk [EY]); advanced yolked 

 oocytes (AY); oocytes with a migratory-nucleus (MN); 

 unovulated hydrated oocytes (HY); ovulated hydrated 

 oocytes (ova); and postovulatory follicles (POP). During 

 the initial stage of oocyte degeneration (known as alpha 

 [a] atresia) the entire oocyte is resorbed by the hyper- 

 trophying granulosa cells of the follicle (Lasker, 1985). 

 Presence of alpha atresia was noted for all histological 

 stages of oocyte development, and the degree of alpha 

 atresia in yolked oocytes was classified as either greater 

 or less than 50% of all yolked oocytes in the ovary. The 

 thickness of the ovarian wall was measured at five or 

 more locations with an ocular micrometer. 



Histological classification 



Based on histological analysis, spawning categories 

 were created for females (Table 1) according to the clas- 



