MAY: EFFECTS ON BAIRDIELLA ICISTIA 



being anesthetized with MS-222 (tricaine 

 methanesulfonate) at 150 ppm. Haydock (1971) 

 found that salmon pituitary glands and PMS were 

 both effective in inducing ovulation in bairdiella. 

 PMS was used here because it had a standardized 

 activity and was more readily available and easier 

 to prepare than salmon pituitaries. The injected 

 fish were checked for ovulation 30 h after injection 

 and at hourly intervals thereafter until ovulation 

 took place (Haydock 1971). In the vast majority of 

 cases, ovulation occurred 30 or 31 h after the hor- 

 mone injection. Spawning bairdiella of the size 

 used in these experiments will jdeld 100,000 or 

 more eggs (Haydock 1971). Male fish remained in 

 a running ripe condition in the laboratory and did 

 not require hormone injections. Bairdiella do not 

 spawn spontaneously in captivity, whether in- 

 jected or not, and gametes must be obtained by 

 stripping. Haydock (1971) demonstrated that eggs 

 must be fertilized 1 or 2 h after ovulation if maxi- 

 mal viability is to be retained. 



Fertilization 



Approximately 1,000 to 3,000 eggs were 

 squeezed from an anesthetized, freshly ovulated 

 female and added to a petri dish containing 75 ml 

 of water of the desired temperature and salinity. 

 When fertilizations under a number of conditions 

 were to be made, eggs were added to all petri 

 dishes before sperm was added. Sperm from a 

 lightly anesthetized male fish was taken up in a 

 pasteur pipette which was immediately filled and 

 flushed with water from a petri dish containing 

 eggs. Eggs and sperm were swirled in the dish for 

 several seconds. This procedure was repeated for 

 every dish, fresh sperm being obtained each time. 

 After cleavage had begun, random samples (usu- 

 ally 100 to 300 eggs) were taken from each petri 

 dish, preserved in 3% Formalin and later ex- 

 amined, and the number cleaving recorded. The 

 percentage of eggs cleaving was taken as the per- 

 centage fertilized. 



Spermatozoan activity was measured in various 

 salinities by placing a drop of sperm under a cover 

 slip, focusing on it with a compound microscope at 

 430 X and adding seawater of the desired salinity. 

 At frequent intervals after hydration, the activity 

 of spermatozoa was rated on an arbitrary scale of 

 to 5, being no activity and 5 being maximal 

 activity. All such tests were conducted at approx- 

 imately 25°C. More than 70 runs were made utiliz- 

 ing spermatozoa from nine fish, each run compris- 



ing between 4 and 15 observations, depending on 

 the duration of activity. 



Incubation 



Developing eggs from the fertilization dishes 

 were counted out by pipette under a dissecting 

 microscope, rinsed with clean water of the test 

 salinity to remove sperm, and transferred to in- 

 cubators. The transfer of eggs was usually com- 

 pleted by the time the blastula stage had been 

 reached, within 3 or 4 h after fertilization. One 

 hundred developing eggs were placed in each in- 

 cubator, and there were two replicate incubators 

 for each experimental treatment. Each incubator 

 (Figure 1) consisted of a 400-ml Pyrex beaker with 

 an insert made from a truncated pol3T)ropylene 

 beaker with its bottom covered by Nitex nylon 

 mesh (350- /i m mesh opening). Three hundred mil- 

 liliters of water were added to each incubator. A 

 slow stream of air bubbles in a centrally 

 positioned glass tube created a flow of water such 

 that eggs which rested on the bottom at low 

 salinities were bathed by a continuous flow of aer- 

 ated water (Figure 1). 



One or two days before each experiment, seawa- 

 ter with a salinity of approximately 60°/oo was 

 made by adding artificial sea salts ("Instant 

 Ocean"; Aquarium Systems, Inc., Wickliffe, Ohio) 

 to HA Millipore-filtered seawater. This solution 

 was filtered through paper (Whatman No. 1) to 

 eliminate a residual cloudiness and then diluted 

 with deionized water to the desired test salinities. 

 Batches of seawater were aerated for several 



Figure 1. — Egg incubator. A) Parafilm cover; B) polyethylene 

 air tube; C) 400-ml Pyrex beaker; D) water line; E) 250-ml 

 IX)lypropylene beaker, cut off at bottom; F) glass chimney; G) 

 Nitex mesh. Arrows indicate direction of water flow. 



