Controlled Maturation of Farmed Salmon Using Salmon GnRHa Implants

In the practice of inducing maturation, there are several different methods that can be used. Historically, gonadotropin hormones (GtH) or pituitary extracts containing GtHs were injected into maturing fish to advance the later stages of reproduction. Human chorionic GtH was also used in this fashion because of its structural similarity to fish GtHs. More recently, mammalian and salmon gonadotropin-releasing hormones (GnRH) have been used to elicit the liberation of GtHs from the pituitary and induce maturation. These hypothalamic hormones are smaller, easier to prepare and are more effective at inducing maturation. Because they are naturally-occurring hormones that use the fishes own internal machinery, they are more reliable and are more potent without harmful side effects to the fish. As well, analogues of both mammalian and salmon GnRH are more potent and degrade slower than natural hormones.

Syndel Labs formulated, tested and developed Ovaprim for use in finfish to induce maturation. Ovaprim was brought on the market in the mid-1980's and has enjoyed international success for inducing maturation in many cultured fish species. Ovaprim is a formulation that contains sGnRH analogue (sGnRHa) and a dopamine antagonist (necessary in many non-salmonid species). The injectable liquid is applied to fish that are currently maturing and have gonads that are in a state of advancing vitellogenisis or spermiogenisis. Ovaprim can induce maturation and coordinate spawning times of maturing populations through liberation of the fishes own GtHs and can induce spawning by as much as one month prior to historic spawning date. This method of inducing maturation is very effective in maturing fish that are close to maturation, but has limitations to induce maturation in immature fish. The best results using Ovaprim are in a population of fish that have begun to spawn. In this way, Ovaprim will compress the spawning season, synchronize spawning times and advance later spawning fish.

Van der Kraak and coworkers (1987) showed that coho salmon could not be induced to ovulate in a period six weeks prior to predicted spawning date, but that fish could be induced to ovulate at four weeks prior to spawning date. However, prolonged release of GnRH analogues through the use of implants has been shown to induce and advance maturation in Atlantic and Pacific salmon^2,. The use of implants is beneficial for a number of reasons:

• analogue levels are sustained for prolonged periods,
• sustained release of sGnRHa results in sustained GtH release²,
• multiple injections cause large fluctuations in plasma levels of hormones and is stressful on fish whereas implants release constant, sustained and stable GtH levels, and
• reduced handling decreases stress and mortality.

Many of the current studies have demonstrated that in use of commercially available GnRH analogues, salmon GnRHa is the most potent. Salmon GnRHa is an analogue of a native peptide and is therefore desirable from a public acceptance standpoint. That is, mammalian GnRH does not appear in teleost fishes whereas salmon GnRH does appear. Salmon GnRHa, the active ingredient in Ovaprim, is proven and accepted as a tool to induce maturation in teleost fishes. Further, salmon GnRHa is ineffective in humans and is therefore safe to use. Therefore, the analogue of choice for incorporation into an implant would be the most potent based on naturally-occurring peptides namely salmon GnRHa.

As world-wide suppliers of Ovaprim, Syndel Labs has been pursued by US and international fish culturists to produce a similar product to Ovaprim but with a sustained release of salmon GnRHa. In response to this demand, Syndel Labs in cooperation with Pacific Aqua Salmon Farming Partners and Aquametrix Research Ltd. of Sidney, B.C. conducted preliminary investigations into the efficacy of a salmon GnRHa-based implant. This work was funded in-house by the collaborating companies.

Atlantic salmon (Salmo salar) historically spawn during the late fall or early winter at local British Columbia latitudes (50^th parallel). This study was conducted with the notion of defining the uppermost limit whereby maturation could be induced. In June of 1996, 20 female and 20 male mature-of-the-year Atlantic salmon were implanted with pellets containing salmon GnRHa. Control fish (maturing cohorts) were not implanted. Experimental fish were held in seawater pens and implanted with another pellet every three weeks until mid-July. When fish were routinely examined in July, male fish were producing copious amounts of milt and female fish had ovulated; 10% of males had motile sperm and 15% of females expressed ova. Other experimental fish matured after the subsequent implantations, but were killed by plankton blooms in August. Hatchery arrangements and logistical troubles precluded incubation of gametes. However, the results of this preliminary trial convinced the collaborators to investigate the use of salmon GnRHa implants for other additional uses.

In follow-up trials in 1997, Atlantic salmon were implanted with pellets of varied amounts of salmon GnRH. In this work, groups of salmon were held in seawater tanks and given one implant of control or four experimental doses. Blood samples were taken from fish every week. Fish that received optimal doses of salmon GnRHa spawned in advance of controls. Milt from these fish was viable and fertilization was complete. Placebo-implanted fish spawned three weeks later than the last experimental fish demonstrating that the implant advances and synchronizes ovulation. Males receiving the implants gave higher volumes of milt for longer periods of time, indicating the efficacy of the implants in both males and females.

Producers often keep large numbers of broodstock, but spawn only early-maturing fish; other later-maturing fish are often not kept until they mature fully. In late October 1997, late maturing fish were separated from cohorts during the peak of spawning season. Female fish (n=36) that were visually greater than three weeks from spawning (ie: green) were each given a salmon GnRHa implant. Control fish were handled, but not implanted. All of the implanted fish produced eggs within 14 days from implantation. Cohort controls continued to mature for three to six weeks. Samples of eggs from implanted fish have developed normally. From these preliminary studies it is clear that sustained-release pellets containing salmon GnRHa can be used to advance spawning date in maturing fish. This advanced maturation can be done well prior to historic spawning date or can be done to coordinate and advance maturation in late spawning fish.

It is noteworthy that every tool in aquaculture has limitations. The best results from using peptide implants come from fish that have a well-defined spawning history. Populations with a smaller spawning window (shorter spawning season) have better results with implants at advancing and synchronizing spawning dates. Implants can be used in photo-manipulated fish will equal efficiency. Again, results vary with individual stocks, but testing salmon GnRH implants in Scotland, Chile, British Columbia and New Brunswick has shown benefit to producers as well as being effective.