FISHERY BULLETIN: VOL. 87, NO. 4, 1989 



MATERIALS AND METHODS 



Uku and onaga caught in 1984-86 and 1985-87, 

 respectively, by commercial fishermen using 

 deep-sea hook-and-line gear were weighed and 

 measured for fork length (FL), and their capture 

 locations were noted. Most were caught in the 

 main Hawaiian Islands and were sold through 

 the Honolulu wholesale fish auction. Following 

 sale of the fish, viscera were extracted by the 

 purchasing agent and refrigerated with an iden- 

 tifying tag. Gonad samples were collected later 

 and preserved at the laboratory either in modi- 

 fied Gilson's fluid (Bagenal and Braum 1968) or 

 Bouin's fluid. 



Sexual maturity of females was evaluated by 

 several methods. First, ovaries were staged 

 macroscopically and given a preliminary matur- 

 ity stage designation (Hilge 1977) (Table 1). To 

 refine and confirm these macroscopic designa- 

 tions, at least one of the following three addi- 

 tional microscopic techniques was used: volu- 

 metric or cork borer subsampling, which is based 

 upon the size and appearance of individual 

 oocytes, or standard histological examination. 

 Hilge's (1977) table was also used to assign a 

 final stage designation to these ovaries. To avoid 

 confusion, prespawning adults prior to vitel- 

 logenesis were classified as stage I immature, 

 and prereproductive individuals as stage I 

 juvenile. 



To determine oocyte size-frequency distribu- 

 tions by volumetric subsampling, uku ovaries 

 preserved in modified Gilson's fluid were ex- 

 amined. After adequate time for dissolution. 



connective tissues were removed, and the re- 

 maining "free" ova were placed in a flask, which 

 was then filled with 200 mL of water. A homo- 

 geneous distribution of ova was obtained by us- 

 ing a magnetic stirrer (Van Dalsen 1977). A 3 

 mL sample was then pipetted onto a gridded 

 petri dish and examined under a binocular dis- 

 secting scope at 50 X. With an ocular micro- 

 meter, 100-200 oocytes were measured along 

 their longest dimension. This method precluded 

 the need to measure oocytes from various sites 

 within the ovary to determine spatial homo- 

 geneity of development. Maturity stages were 

 assigned based upon the largest oocyte mode and 

 the degree of oocyte transparency (Table 1). 



Subsamples of ovaries of both species pre- 

 served in Bouin's fluid were taken from the 

 anterior portion with a cork borer and examined 

 under a binocular dissecting microscope at 50 x . 

 The average diameter of the largest oocyte mode 

 was determined, and the percentage of each 

 maturity stage present was noted. Oocyte di- 

 ameter frequency plots were constructed for uku 

 ovaries in various stages of development and 

 compared with similar plots constructed by us- 

 ing oocyte diameter data obtained by the vol- 

 umetric method. 



For histological examination, some ovaries of 

 both species representing various visually iden- 

 tifiable maturity stages were transferred from 

 Bouin's fluid to 70% ethanol. Portions of ovaries 

 from 28 uku and 22 onaga caught at various 

 times of the year were embedded in paraffin, 

 sectioned at 5 ixm, stained with hematoxylin, 

 and counterstained with eosin. Each was as- 



Table 1 . — Ovary developmental stage designations used for study of the reproductive 

 cycle of uku and onaga. Designations are adapted from Hilge (1977). 



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