Photoperiod and gonadotropin control of the
reproductive cycle in the female rainbow trout

Bernard Breton, Roland Billard

► To cite this version:

Bernard Breton, Roland Billard. Photoperiod and gonadotropin control of the reproductive cycle in the female rainbow trout. Current trends in comparative endocrinology, Hong-Kong University Press, 1985, 962-209-039-7. hal-01601076

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# Photoperiod and Gonadotropin Control of the Reproductive Cycle in the Female Rainbow Trout

B. Breton, Y. Zohar and R. Billard

Laboratoire de Physiologie des Poissons, I.N.R.A., Rennes, France

Important changes in GtH function are found to be associated with three critical periods during the salmonid annual reproductive cycle.¹⁻³ In spring a rise in plasma GtH corresponds to a possible pulsatile mode of GtH secretion preceding the beginning of exogenous vitellogenesis. In June, no GtH pulsations are detected but then GtH levels increase slightly in late summer and early autumn, concomitant with the rapid growth of oocytes. During this period GtH secretions are again pulsatile. Finally, GtH secretion increase during oocyte maturation. The aim of the present study was to try to determine to what extent the events depend on the photoperiod, and to explain the mechanism by which shortened daylengths stimulate gametogenesis.⁵

## Materials and methods

Groups of female rainbow trout, which had spawned normally during the first half of December, were placed in a recirculating water system, and maintained either under natural light or experimental photoperiods. The latter consisted of a 6 months decreasing daylength regime (16L:8D to 8L:16D), initiated from December 21, or from 3, 4, 5 or 6 months after this date. Experiments were carried out at 10 and 15°C, and the following parameters were studied: (i) the evolution of oogenesis, evaluated on the basis of the gonadosomatic index (GSI) and histological observations; (ii) GtH plasma levels in bimonthly blood samples taken from the same 10 fish in each group; (iii) pituitary responsiveness to LHRH stimulation,⁶ at the beginning of the experiment and 15 days before and after the start of the decreasing light period, and until it reached its minimum value; and (iv), gonadal sensitivity to GtH stimulation assessed by measuring the ability of GtH to induce *in vivo* estradiol (E₂) secretion,⁴ using the same

sampling frequency as in (iii). Results were analyzed using the couple method after logarithmic transformation of results and variance analysis.

## Evolution of gametogenesis

Ovulation first occured some 6 months after the start of the experimental photoperiodic regime. Generally there was no difference between fish kept at 15 and 10°C, except in those fish that had been maintained for 3 months at 10°C under increasing natural daylengths before being placed under experimental photoperiods. In these fish ovulation was delayed by 20 days as compared with those kept at 15°C under identical photoperiodic conditions. Immediately after ovulation oocytes at the endogenous vitellogenesis stage were identified in all groups. All fish were in this stage in early February. Exogenous vitellogenesis was recorded in all experimental photoperiod groups 1.5 months after the start of their exposure to decreasing light (Fig. 1) then the GSI increased rapidly, except in the group that had first experienced 3 months of natural increasing daylengths before the decreasing regime began. In the latter, and also in controls, exogenous vitellogenesis occurred at first without any significant increase in the GSI, then this was followed by a rapid GSI increase.

## Plasma gonadotropin

GtH levels were measured in control and experimental groups from blood samples taken at the same time of day. The high post-ovulatory GtH levels decreased to a basal level within a few weeks, and were significantly higher in fish that had been subjected early on to decreasing light (Fig. 2). During the rest of the experiment

*Current Trends in Comparative Endocrinology* (eds. B. Lofts & W.N. Holmes). Hong Kong, Hong Kong University Press, 1985.
<figure><img src="image_2.png" /><figcaption>Fig. 1. Evolution of gametogenesis.</figcaption></figure>

<figure><img src="image_3.png" /><figcaption>Fig. 2. Profiles of plasma GtH section under different photoperiodic regimes: (A) decreasing light from December 21; (B) decreasing light from February 2; and (C) decreasing light from April 24.</figcaption></figure>

significant episodic rises in GtH were recorded in all groups; on February 2, in the fish exposed to decreasing daylengths from December 21 (basal level around 4 ng/ml); on March 3, in the group where decreasing light was applied from February 2 (basal level around 3 ng/ml). These rises always preceded exogenous vitellogenesis by about one month. In the group that had been kept under 3

months of natural daylengths before the artificial photoperiod regime began, 2 episodic increases of plasma GtH were seen, one in early March (when photoperiods had reached 10L:14D), and a second one in early August just prior to rapid oocyte growth. In the control group a single period of increasing plasma GtH levels was observed in late March. All fish exhibited a rise in plasma GtH levels during the breeding period.

## Pituitary gonadotropin levels

These decreased in controls from over 20 µg/mg fresh tissue in December, to a minimal level of 5 µg in May. The decrease occurred progressively, and levels determined in December and February were not significantly different. A similar decrease occurred in fish kept under modified light regimes, but the GtH pituitary content rose again at the beginning of May while daylength was decreasing, and was higher under 11h than 14h of light. Temperature did not induce significant differences in the pituitary GtH content. There was also no correlation between plasma and pituitary GtH concentration at any time under increasing or decreasing photoperiods.

## Pituitary responsiveness to LHRH stimulation

The index of pituitary responsiveness was expressed as the difference between the total amount of GtH secreted after an LHRH injection (1 µg/kg body wt) and after a saline injection during the 20 min following each injection. In fish maintained under natural photoperiods, it decreased from the time of ovulation to a minimum in May, then increased again at the end of June, when the photoperiods had reached 16h of light. The response subsequently remained positive for the rest of the year. A transfer to the artificial light regime induced a rapid disappearance of pituitary responsiveness as early as February (14L:10D), and remained negative during March. A positive response was recorded in early May in the groups receiving 11h and 14h daily illumination, and was significantly higher (P < 0.05) under the shortest daylength. Under modified light regimes and natural conditions, increasing GtH pituitary content and
pituitary responsiveness to LHRH occurred at the same time.

## Gonad responsiveness to GtH stimulation

Saline injections never stimulated E₂ secretion. After GtH treatment the response (expressed by a significant increase in plasma E₂ levels, 3 or 6h after a single intravenous injection of 5 µg/kg body of GtH) showed that in controls, as long as fishes were kept under increasing light, GtH did not stimulate E₂. However, controls became responsive in late June, and by mid September, under a 12h decreasing daylength, all fish responded positively. In all experimental groups GtH never stimulated *in vivo* E₂ secretion until the end of March, even if the fish had already been subjected to decreasing light for 2 or 3 months. However from May onwards, under 14h of daylength, 12% of the fishes because responsive to GtH treatment, their number increasing with decreasing light conditions.

## Discussion

The results show that following ovulation, there was a rapid transition from a previtellogenetic condition to a vitellogenetic stage, which appeared to be independent of the photoperiodic regime. This endogenous vitellogenesis continued to develop independent of the environmental component. Exogenous vitellogenesis appeared under increasing light conditions in the controls, and decreasing light in all other groups. However, rapid gonad growth never took place before the decreasing photoperiod had reached 14h of light. Thus, exogenous vitellogenesis appeared more strictly correlated with decreasing photoperiods.⁵ The duration of rapid ovarian growth was similar in all groups, whereas the slow ovarian growth phase appeared to depend upon the date on which the experimental light regime began. Thus, it was absent when decreasing photoperiods were applied before February, but when they were started after a 3 months exposure to the naturally increasing daylengths, a distinction was seen between a slow and a rapid gonad growth during exogenous vitellogenesis. Fishes transferred into decreasing light

conditions in late April and also controls, showed no significant changes in their GSI in the following 2 and 3 months after the start of exogenous vitellogenesis. The beginning of this latter process was, in all groups, preceded by a transient surge of GtH secretion in early spring, without a concomitant indication of pituitary LHRH responsiveness. This rise of GtH probably corresponds to the episodic, high amplitude elevations (pulses) in blood GtH levels observed under natural conditions, when oocytes undergoing early exogenous vitellogenesis are present in the ovaries.³ Same spring rises also occurred before the first detection of exogenous vitellogenesis under decreasing light, when there was no distinction between a slow and a rapid growth of the ovaries. Thus, in spring there was always a transient GtH rise, which, depending on the group, occurred from early February to the end of March, independent of the photoperiod, and possibly corresponds to the gonadotropic initiation of exogenous vitellogenesis.

The sensitivity of our *in vivo* method did not allow any detection of an ovarian response to GtH stimulation before the end of June under natural conditions although *in vitro* E₂ synthesis increase in May⁷ and could be stimulated by GtH as early as March (Zohar, unpublished). This might reflect either an inhibitory action on synthesis or secretion of E₂ *in vivo*, or a rapid clearance of E₂ during the initiation of exogenous vitellogenesis. Under modified photoperiods, following 3 months of apparent ovarian refractoriness to GtH, it appeared that GtH-stimulated E₂ secretion may depend on decreasing photoperiods. The pituitary responsiveness to LHRH stimulation in controls decreased from ovulation to the end of June, when it increased again under decreasing light to reach its maximum during spawning.⁶ Under modified photoperiods it did not appear with the decreasing light until May, then, from this point onwards it became positive again under decreasing daylengths, concomitant to the increase of pituitary GtH content. This allows the supposition that pituitary responsiveness to LHRH may be correlated with an active synthesis of pituitary GtH. There was no detectable *in vivo* pituitary responsiveness at the time of the spring rise in plasma GtH, which suggests that a possible GtH-induced exogenous vitellogenesis initiation
could be controlled either by a gonadal steroid
feedback, or by a central inhibitory mechanism,
which has already been suggested to play a role in
the GtH control of evulation in goldfish,⁸ and
which would not depend on photoperiod in the
trout. Later, decreasing photoperiods would lead
to the supposed pulsatile mode of GtH secretion
stimulated by GnRH.

Acknowledgements

This work was supported by a grant C.N.R.S. -
C.N.E.X.O. No. 3562. We are very grateful to
Mrs Reinaud and Sambroni for their helpful
technical assistance.

References

¹ Breton, B., Fostier, A., Zohar, Y., Le Bail, P.Y. & Billard, R. (1982). *Gen. Comp. Endocrinol*. In press.

² Billard, R., Breton, B., Fostier, A., Jalabert, B. & Weil, C. (1978). In *Comparative Endocrinology* (eds. Gaillard, P.J. & Boer), pp. 37–47. Elsevier/North Holland Biomedical Press, Amsterdam.

³ Zohar, Y., Breton, B. & Billard, R. (1981). *Gen. Comp. Endocrinol*. In press.

⁴ Fostier, A., Breton, B. & Jalabert, B. (1979). *Ann. Endocrinol. 40*, 83-84.

⁵ Henderson, N.E. (1963). *J. Fish. Res. Board Can.* **20**, 854-97.

⁶ Weil, C. (1981). Thesé doctorat d'Etat — Université de Paris VI.

⁷ Van Bohemen, Ch. G. & Lambert, J.G.D. (1981). *Gen. Comp. Endocrinol.* **45**, 105-14.

⁸ Peter, R.E. & Paulencu, C.R. (1980). *Neuroendocrinology* **31**, 133-41.
Current Trends in
Comparative Endocrinology

Editors

B. Lofts and W. N. Holmes

Volume Two

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Hong Kong University Press

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