698 



Fishery Bulletin 92|4), 1994 



The weakfish, Cynoscion regalis, is a sciaenid 

 which spawns in bays and estuaries from North Caro- 

 lina to Long Island, New York, during the spring and 

 early summer (Welsh and Breder, 1923; Mercer, 

 1983). Merriner (1976) noted a change in the colora- 

 tion of the sonic muscles in male weakfish that par- 

 alleled the changes in testis condition during the 

 course of the year. 



The purpose of the present study was to determine 

 whether the condition of the sonic muscles of male 

 weakfish changes seasonally. In particular, this study 

 was designed to determine the extent of change, if 

 any, in the morphometries of the sonic muscles over 

 the course of the spawning season and to observe 

 these variations in relation to changes in testis con- 

 dition and plasma androgen levels. 



Materials and methods 



Sample collection 



Male weakfish were sampled near the mouth of the 

 Delaware Bay (lat. 38°50.30'N, long. 07512. 92'W) 

 and roughly 40 km north of this (39°11.98'N, 

 075°23.20'W). Field collections were made from May 

 through September in 1990 and from April through 

 September in 1991. Specimens were collected by 

 means of anchored or drifting gill nets, hook and line, 

 and otter trawl. 



Immediately after capture of the fish, blood 

 samples were taken with heparinized syringes from 

 the hemal canal, posterior to the anal fin. The blood 

 was then placed in heparinized microcentrifuge tubes 

 and stored on ice. Samples were centrifuged at 2,000 

 x g, and the supernatant was removed and frozen at 

 -80°C for determination of plasma testosterone and 

 11-ketotestosterone levels via radioimmunoassay 

 (RIA). In 1991, blood sampling was preceded by milt 

 collection to determine the number of ripe specimens. 

 Any drumming behavior was also noted. 



Autopsies of the specimens provided total length 

 (TL), total weight (TW), testis weight, and morpho- 

 metric measurements of the sonic muscles. Testis 

 weight and total weight were used to calculate a 

 gonadosomatic index (GSI=|total testis weight/total 

 weightl x 100). Sonic muscle weight, width (anterior- 

 posterior axis of the muscle), length (dorso-ventral 

 axis of the muscle), and maximal thickness (cross- 

 section of the muscle) were measured for both the 

 right and left sonic muscles. Orientation of sonic 

 muscle width and length measurements is based on 

 the dorso-ventral orientation of the muscle fibers 

 (Ono and Poss, 1982). The sonic muscle-somatic in- 

 dex (SMSI) was calculated as SMSI = {total sonic 



muscle weight/total weight x 100), and the results 

 were expressed as a percentage of TW. Indices for 

 sonic muscle width (SMWI), length (SMLI) and thick- 

 ness (SMTI) were calculated as the mean of the mea- 

 surement for the right and left sonic muscles/total 

 length x 100 and were expressed as a percentage of 

 TL. The color of the sonic muscles was also noted. 



Radioimmunoassays 



Testosterone was measured by direct radioimmu- 

 noassay. Five uL aliquots of serum were placed in 2- 

 mL conical glass tubes (methanol rinsed) and diluted 

 to a total volume of 50 uL with borate buffer. Di- 

 luted samples were incubated at 60°C for one hour 

 to dissociate the steroid from binding proteins. Stan- 

 dard solutions were prepared by dissolving crystal- 

 line testosterone ( Sigma Chemical Co., St. Louis, MO) 

 in absolute ethanol. Working standards (5, 10, 25, 

 50, 100, 250, 500 pg testosterone 5 uL 1 ) were pre- 

 pared in ethanol, dried to zero volume at 45°C under 

 vacuum, and reconstituted in 50 uL of borate buffer. 

 Standards and sample tubes were incubated with 100 

 uL (approximately 4,000 cpm) of dilute trace ( 1,2,6,7" 

 3 H testosterone, cat. #NET-370, New England 

 Nuclear Corporation) and 100 uL of reconstituted 

 antiserum (Wein Laboratories, Succasunna, NJ) 

 overnight at room temperature. Total counts were 

 estimated by using vials containing 100 uL of dilute 

 trace and 100 uL of saturated ammonium sulfate. 

 Triplicate standard and serum samples incubated 

 without the addition of antiserum were used to cal- 

 culate nonspecific binding. Bound steroids were pre- 

 cipitated by adding 250 uL of saturated ammonium 

 sulfate to each tube. The vials were centrifuged and 

 400 uL of the supernatant were removed and placed 

 in counting vials along with 6 mL of scintillation cock- 

 tail. All tubes were shaken for 25 minutes, allowed 

 to sit for at least one hour, and counted for 3-10 min- 

 utes in a liquid scintillation counter. 



Testosterone measurements were assumed to es- 

 timate total testosterone levels, as the plasma was 

 incubated at 60C for one hour to dissociate any bind- 

 ing proteins in the plasma. The 11-ketotestosterone 

 assay (Woods and Sullivan, 1993 ) measured only free, 

 or unbound, steroid, as the protocol includes tripli- 

 cate ethyl-ether extraction of unbound steroids from 

 the plasma. 



Cross-reactivities of the testosterone antiserum 

 were >50% for 5a-dihydrotestosterone and 5-1-test- 

 osterone, approximately 18% and 12.5% for 5a- 

 androsten-3p,17P-diol and 5-5-androsten-3p,17P-diol, 

 respectively, and <5% for all other steroids tested 

 (Wein Laboratories, Inc.). Tritiated 11-ketotestoster- 

 one and 11-ketotestosterone antiserum were gifts 



