Ž .
barus clarkii, by acting to stimulate release of a gonad-stimulating hormone GSH from the brain and thoracic ganglia; GSH is an abstract entity, which has not been identified
nor measured directly. Ž
. Our current knowledge on regulation of GSH and gonad-inhibiting hormone GIH
Ž .
release was presented by Fingerman 1997b . GSH release is stimulated by 5-HT and Ž
. red pigment-concentrating hormone RPCH ; GSH release is inhibited by dopamine
Ž .
Ž .
DA and methionine enkephalin Met-ENK ; GIH release is stimulated by Met-ENK and DA, and GIH release inhibition has not been reported.
Serotonin also stimulates the release of other neurohormones, including the crus- Ž
. Ž
. tacean hyperglycemic hormone CHH , red pigment-dispersing hormone RPDH , neu-
Ž .
Ž .
rodepressing hormone NDH and molt-inhibiting hormone MIH; Sarojini et al., 1994 . Ž
. Ž
Methyl farnesoate MF is a factor that stimulates gonadal maturation Laufer et al., .
1993 and its synthesis is inhibited by 5-HT; on the other hand, RPCH stimulates MF Ž
. synthesis by the mandibular organ Fingerman, 1997b . Other roles played by 5-HT are
migration of the proximal retinal pigment, pericardial organ neurohormone, stomatogas- tric ganglion neuromodulator or neurohormone, behavioral responses, osmoregulation,
Ž .
and mechanoreception Fingerman et al., 1994 . As an effort to develop alternatives for the commercial reproduction of penaeid
shrimp, this study was conducted to evaluate the effect of two doses of 5-HT on P. Õ
annamei maturation and spawning quality, compared to unilateral eyestalk ablation.
2. Material and methods
2.1. Broodstock Adult P. Õannamei with a mean weight of 56 g for females and 40 g for males, were
obtained by trawling off Golfo de Panama, on the Pacific coast. Animals were
´
transported in oxygenated seawater to the Larvae Production Center of Cocle shrimp
´
farm, Panama. Selection was done based on size, stage of ovarian maturation, and
´
healthy appearance. 2.2. Experimental conditions
Ž
2
. Maturation system consisted of three concrete tanks 26 m reach , with a water
depth of 0.35 m and total daily replacement of 200. Light intensity was approximately 2 mE m
y2
s
y1
, with a photoperiod of 8 h light: 16 h dark; these conditions are routinely used in the shrimp farm. Water temperature was 29.5 0.88C, salinity between 30 and
34 ppt. Animals were fed at 21 body weightrday with fresh frozen squids, blood- worms and oysters at a ratio of 1:1:1, respectively.
Molting and reproduction are major metabolic processes in decapod Crustacea Ž
. Adiyodi and Adiyodi, 1970 ; therefore, in penaeid shrimp ovarian maturation and
Ž .
spawning occur during the intermolt Browdy, 1992 . The molt cycle duration of adult Ž
. Penaeus is 11–14 days Robertson et al., 1987 ; at postmolt females are weak and at
Ž .
premolt maturation is delayed. During the adaptation period 2 weeks to the experimen-
tal environment, molting stages of females were determined by uropod analysis, based Ž
. Ž
. on the technique described by Robertson et al. 1987 . Intermolt females 2–3 days
were randomly distributed in a block design, integrated by three tanks with four treatments each, and 12 females per treatment-tank, for a total number of 36 females per
treatment. Each female was individually eyestalk tagged; tank density was adjusted to 5 animalsrm
2
using non-treated animals, with 1:1 sex ratio. Treatment A consisted of unilateral eyestalk ablation by cutting the right eyestalk
Ž with red-heated scissors; animals in treatment B received three injections days 1, 11 and
. Ž
.
y1
21 of 5-HT creatinine sulfate Sigma, St. Louis, MO, USA at 15 mg g body weight
Ž .
Ž .
b.w., Sarojini et al., 1995 . Treatment C received three injections days 1, 11 and 21 of 5-HT at 50 mg g
y1
b.w. Treatment D served as control receiving three injections of Ž
. sterile vehicle solution
NaCl 0.85 , at the same experimental days. Volume of injection was 0.07 ml per 56 g; the study was undertaken for 46 days.
Ovarian maturation was evaluated by external observation of ovarian size and color Ž
. Ž
. as described by King 1948 and Yano et al. 1988 with slight modifications:
Stage I. The ovary is transparent with no distinguishable outline. Stage II. The ovary is visible as a thin opaque line along the dorsal central axis.
Stage III. The ovary is visible as a thick and yellow band. Stage IV. The ovary is turgid, broad and dark orange. Mating and spawning are
imminent.
Quality of spawns was evaluated in terms of number of eggs per spawn, percentage of fertilized eggs, hatching rate, and number of nauplii per spawn. Mature and mated
Ž .
females were selected and placed in individual spawning tanks 80 l ; 10 h after spawning, the eggs were washed and concentrated in a 10-l container, three 1-ml
samples were taken to asses total number of eggs, additional samples were taken for a microscopic investigation of the proportion of normally embryonal eggs vs. non-em-
Ž .
Ž .
bryonal eggs Primavera and Posadas, 1981 . Eggs were treated with formalin 37 Ž
.
y1 y1
and iodine 1 at 0.26 ml l and 0.20 ml l
, respectively; then moved to hatching Ž
. containers 60 l at salinity of 30 ppt. Thirty-four hours after spawning, nauplii were
collected and concentrated in 15 l containers; three samples of 1 ml were randomly taken to count number of nauplii. Hatching rate was estimated based on number of eggs
and nauplii produced per spawn.
2.3. Statistical analysis Ž
. Experimental data were analyzed with Statgraphics plus version 7.0 , for one way
Ž .
classification analysis of variance AOV and Duncan’s multiple range procedure. Prior Ž
to analysis, data on percentages were transformed using squared root of arcsine Bray et .
al., 1990 , to make the variance independent of the mean; alpha levels for all tests were Ž
. set at 0.05. Untransformed data are presented as mean standard error s.e. .
3. Results
