G. Van Der Kraak

Exposure to bleached kraft pulp mill effluent disrupts the pituitary-gonadal axis of white sucker at multiple sites

Our recent studies have demonstrated reproductive problems in white sucker (Catostomus commersoni) exposed to bleached kraft pulp mill effluent (BKME) at Jackfish Bay on Lake Superior. These fish exhibit delayed sexual maturity, reduced gonadal size, reduced secondary sexual characteristics, and circulating steroid levels depressed relative to those of reference populations. The present studies were designed to evaluate sites in the pituitary-gonadal axis of prespawning white sucker affected by BKME exposure. At the time of entry to the spawning stream, plasma levels of immunoreactive gonadotropin (GtH)-II (LH-type GtH) in male and female white sucker were 30- and 50-fold lower, respectively, than the levels in fish from a reference site. A single intraperitoneal injection of D-Arg6, Pro9N-Et sGnRH (sGnRH-A, 0.1 mg/kg) increased plasma GtH levels in male and female fish at both sites, although the magnitude of the response was greatly reduced in BKME-exposed fish. Fish at the BKME site did not ovulate in response to sGnRH-A, while 10 of 10 fish from the reference site ovulated within 6 hr. Plasma 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17,20 beta-P) levels were depressed in BKME-exposed fish and unlike fish at the reference site, failed to increase in response to sGnRH-A. Testosterone levels in both sexes and 11-ketostestosterone levels in males were elevated in fish from the reference site but were not further increased by GnRH treatment. In contrast, BKME-exposed fish exhibit a transitory increase in testosterone levels in response to the GnRH analog. In vitro incubations of ovarian follicles obtained from fish at the BKME site revealed depressed basal secretion of testosterone and 17,20 beta-P and reduced responsiveness to the GtH analog human chorionic gonadotropin and to forskolin, a direct activator of adenylate cyclase. By comparison, ovarian follicles from fish collected at BKME and reference sites produced similar levels of prostaglandin E basally and in response to a phorbol ester and calcium ionophore A23187, suggesting that BKME effects on ovarian function are selective and do not reflect a general impairment of ovarian function. BKME-exposed fish had plasma levels of testosterone glucuronide proportionately lower than those of reference fish, suggesting that there are site differences in the peripheral metabolism of steroids. These studies demonstrate that BKME exposure affects reproduction by acting at multiple sites in the pituitary-gonadal axis.

Changes in hepatic mixed-function oxygenase (MFO) activity, plasma steroid levels and age at maturity of a white sucker (Catostomus commersoni) population exposed to bleached kraft pulp mill effluent

Abstract The impacts of bleached kraft mill effluent (BKME) on a white sucker (Catostomus commersoni) population were examined during May, July and August 1989, and compared with two reference sites. At the time of this study, the effluent received only primary treatment. BKME-exposed white sucker exhibited increased liversomatic indices and elevated mixed-function oxygenase (MFO) activity in both July and August. They also showed lower gonadosomatic indices and an increased age to maturity. The females contained fewer eggs at maturity, while the males had reduced development of secondary sexual characteristics. These fish also had severe reductions in plasma steroid levels throughout the year, including testosterone, and 17α,20β-dihydroxyprogesterone in both sexes, as well as 11-ketotestosterone in males and 17β-estradiol in females. BKME-exposed white sucker were shorter, older and had decreased growth rates compared to those at the reference sites. These fish also exhibited an increased condition factor, yet showed decreased visceral lipid stores. Relative to those at the reference sites, the stomach contents of the BKME-exposed fish revealed reduced numbers of organisms per gut, reduced taxa per gut and an increased number of empty stomachs. The decreased energetic commitment to reproduction, along with the increased condition factor, suggested a disruption in metabolic capability and altered energy allocation in fish exposed to BKME.

Effect of holding temperature on ovulation, egg fertility, plasma levels of reproductive hormones and in vitro ovarian steroidogenesis in the rainbow trout Oncorhynchus mykiss

Abstract Sexually mature female rainbow trout were held at temperatures ranging from 9–21 °C for up to 3 months prior to the natural time of ovulation, in experiments conducted over three different spawning seasons. The majority of fish held at 9 and 12 °C ovulated, variable numbers ovulated at 15 and 18 °C and only one fish ovulated at 21 °C. Egg production was similar at 9, 12 and 15 °C, significantly lower at 18 °C and near zero at 21 °C. Egg survival to the eyed stage after incubation at 11 °C was similar at 9, 12 and 15 °C, and nil at 18 and 21 °C. Histological examination of oocytes from fish held at 12, 15 or 18 °C for 1 or 2 months showed no evidence of gonadal atresia. Plasma levels of gonadotropin (GtH) were measured in samples taken 1, 2 and 3 months after introduction to temperatures of 9, 12, 15, 18 or 21 °C, and showed no differences between temperatures at any time. Plasma levels of testosterone (T) and 17β-oestradiol (E 2 ) were similarly unaffected. Repeat measurement of T and E 2 in another spawning season also showed that holding temperature had no effect on plasma steroid levels. In contrast, in vitro basal steroidogenesis by isolated ovarian follicles was generally lower at 18 °C than at 12 or 15 °C. Follicles from fish held at 18 °C for 2 months did not retain responsiveness to stimulation with steroid precursors or GtH, whereas those from fish held at 12 and 15 °C did. The results indicate that elevated autumn holding temperatures have a deleterious effect on ovulation, egg production and fertility but have equivocal effects on endocrine parameters associated with vitellogenesis. This suggests that the effects are exercised on processes associated with final maturation and ovulation rather than vitellogenesis.

Exposure to high temperature influences the behaviour, physiology, and survival of sockeye salmon during spawning migration

Since 1996, some populations of Fraser River sockeye salmon (Oncorhynchus nerkaWalbaum in Artedi, 1792) have begun spawning migrations weeks earlier than normal, and most perish en route as a result. We suspect that a high midsummer river temperature is the principal cause of mortality. We intercepted 100 sockeye during normal migration near a spawning stream and measured somatic energy and aspects of plasma biochemistry. Fish were then held at either 10 or 18 8C for 24 days. Before release, fish were biopsied again and implanted with acoustic transmitters. A group of bi- opsied but untreated control salmon were released at the same time. Sixty-two percent (8 of 13) of control salmon and 68% (21 of 31) of 10 8C salmon reached spawning areas. The 18 8C-treated fish were half as successful (35%; 6 of 17). During the holding period, mortality was 2 times higher and levels of Parvicapsula minibicornis(Kent, Whitaker and Dawe, 1997) infection were higher in the 18 8C-treated group than in the 10 8C-treated group. The only physiological dif- ference between treatments was a change in gill Na + ,K + -ATPase activity. This drop correlated negatively with travel times for the 18 8C-treated males. Reproductive-hormone levels and stress measures did not differ between treatments but showed significant correlations with individual travel times.

An overview of recent studies on the potential of pulp‐mill effluents to alter reproductive parameters in fish

In the early 1990s, many Canadian pulp and paper mills implemented process changes to comply with new regulations that came into effect in 1993. These regulations placed stricter guidelines on a number of parameters in effluent discharges, including limits on acute toxicity, on the discharges of suspended solids, and on biochemical oxygen demand. To meet these new regulations, many of the older Canadian pulp and paper mills had to install secondary treatment systems. The investment by the Canadian pulp and paper industry was in excess of

Developing a Mechanistic Understanding of Fish Migrations by Linking Telemetry with Physiology, Behavior, Genomics and Experimental Biology: An Interdisciplinary Case Study on Adult Fraser River Sockeye Salmon

5 billion, and the implementation of the new regulations and the process changes took several years. The new regulations were an extension of regulations designed in the early 1970s and were not designed specifically to address the reproductive responses recently reported in fish collected downstream of mills in Scandinavia and North America. This report describes a series of projects conducted between 1991 and 1996 to evaluate the effectiveness of the new regulations to address the issue of reproductive responses in fish associated with exposure to pulp-mill effluents. These studies have shown that the existing short-term bioassays do not adequately predict the potential of effluents to affect reproduction in wild fish. Laboratory testing using fathead minnows exposed over a full life cycle confirmed depression in sex steroid production, delay in sexual maturity, reduced egg production, and changes in secondary sex characteristics documented at some sites. Our studies demonstrated that both steroid hormone changes and induction of liver detoxification enzymes take place quickly. While short-term exposures can predict the potential of some effluents to impact steroid hormone production, there is no readily available assay that can be widely applied. In the absence of a usable and transferable laboratory bioassay, field collections were conducted at a number of sites. Generalizations are not possible at this time, but impacts have been seen at a variety of sites, and partial recovery has been documented at five sites in North America following various process and waste treatment changes. Data gaps and critical research areas are identified.

An investigation into the role of prostaglandins in zebrafish oocyte maturation and ovulation

Abstract Fish migration represents one of the most complex and intriguing biological phenomena in the animal kingdom. How do fish migrate such vast distances? What are the costs and benefits of migration? Some of these fundamental questions have been addressed through the use of telemetry. However, telemetry alone has not and will not yield a complete understanding of the migration biology of fish a or provide solutions to problems such as identifying physical barriers to migration or understanding potential impacts of climate change. Telemetry can be coupled with other tools and techniques to yield new insights into animal biology. Using Fraser River sockeye salmon (Oncorhynchus nerka) as a model, we summarize the advances that we have made in understanding salmonid migration biology through the integration of disciplines (i.e., interdisciplinary research) including physiology, behavior, functional genomics, and experimental biology. We also discuss opportunities for using large-scale telemetry arrays an...

Reproductive and stress hormone levels in goldfish (Carassius auratus) exposed to oil sands process-affected water

This study explored the potential for ovarian-derived prostaglandins (PGs) to be involved in the regulation of oocyte maturation and ovulation in zebrafish. It was demonstrated that cultured vitellogenic follicles have the capacity to produce prostaglandin E(2) (PGE(2)) and PGF(2alpha) in response to arachidonic acid (AA) in a concentration-dependent manner, and that AA stimulates the in vitro production of 17beta-estradiol (E(2)). The production of AA-stimulated PGF(2alpha) was significantly reduced by treatment with the non-selective cyclooxygenase (COX) inhibitor, indomethacin (INDO). Treatment of full-grown follicles with AA did not induce oocyte maturation as assessed by germinal vesicle breakdown, but INDO significantly decreased the rate of spontaneous maturation. Using Real-Time PCR, it was shown that follicles of different developmental size classes (primary growth and pre-vitellogenic, early-vitellogenic, and mid- to full-grown vitellogenic) express enzymes that release (cytosolic phospholipase A(2) (cPLA(2)); phospholipase Cgamma1) or metabolize (COX-1, COX-2, and prostaglandin synthase-2) AA to PG metabolites. The expression of cPLA(2) was found to be significantly greater in full-grown follicles compared to follicles of the pre- and early-vitellogenic stages. In vivo studies demonstrated that breeding groups of zebrafish exposed to 100 microg/L INDO exhibited reduced spawning rates and clutch sizes compared with control and 1 microg/L INDO exposed fish. In other studies, it was shown that naturally spawning groups of females exhibit increased ovarian levels of PGF(2alpha), E(2), and 17alpha,20beta-dihydroxy-4-pregnen-3-one (a maturation-inducing hormone in zebrafish) near the time of ovulation compared with non-breeding females. Collectively, these experiments indicate that the AA pathway in zebrafish ovaries is involved in the regulation of oocyte maturation and ovulation and a non-selective inhibitor of COX disrupts these processes.

Seasonal migrations and reproductive patterns in the lake sturgeon, Acipenser fulvescens, in the vicinity of hydroelectric stations in northern Ontario

Athabasca oil sands mining in northern Alberta produces process-affected waters that are characterized by the presence of naphthenic acids, polycyclic aromatic hydrocarbons, and high salinity. The purpose of this study was to examine the impact of these process-affected waters on reproductive and stress related endpoints in mature goldfish, Carassius auratus. In two separate studies, testosterone and 17beta-estradiol levels in the plasma were significantly reduced in both male and female goldfish caged for 19 days in process-affected waters relative to controls. This effect was most pronounced in goldfish caged at a site containing mature fine tailing and tailings pond water (P5). Ovarian and testicular tissues from fish in the caging studies were incubated in vitro to evaluate potential differences in basal steroid production levels and responsiveness to human chorionic gonadotropin (hCG). Basal levels of testosterone were reduced significantly in males and females from P5 compared with the control pond (P1) demonstrating that the gonads from exposed fish had a diminished steroidogenic capacity. Gonadal tissues of fish from all ponds responded similarly to hCG suggesting that the steroid biosynthetic pathway remained functionally intact. Plasma cortisol levels were significantly higher in male goldfish caged in a pond containing mature fine tailings and capped with uncontaminated water (P3) and in P5 compared with P1. Collectively, these studies suggest that waste products of oil sands mining have the potential to disrupt the normal endocrine functioning in exposed fish through alterations to both reproductive and glucocorticoid hormone biosynthesis. In additional laboratory studies, exposure of goldfish to a naphthenic acid extract for 7 days failed to replicate the effects of processes-affected waters on plasma steroid levels and the causative agent(s) responsible for the effects on steroid biosynthesis remains to be identified.

Exposure to bleached kraft pulp mill effluent reduces the steroid biosynthetic capacity of white sucker ovarian follicles

This study was conducted in order to evaluate seasonal migratory behaviour and reproductive pattern of lake sturgeon in a confined region of the Mattagami River system in northern Ontario where river flow is regulated by hydroelectric works. Radio tracking and the systematic sampling of lake sturgeon using gill nets indicated that the distribution of fish throughout the study site varied on a seasonal basis. This distribution was related to the migration of individuals to potential spawning sites in the spring, a post-spawning dispersal to feeding areas and late summer migration to an area of concentration on the Groundhog River which is a tributary of the Mattagami River. There was a high proportion of fish (about 50%), within the size range of reproductively active fish, found in the vicinity of suitable spawning habitat during early May. Measurement of the gonadosomatic index (GSI) and plasma sex steroid hormone levels revealed a divergent pattern of reproductive development between the sexes. Female sturgeon exhibited a prolonged period of ovarian regression following spawning. Resumption of ovarian development was not evident until September and was characterized by an increased GSI and plasma levels of testosterone and 17β-estradiol. In contrast, male lake sturgeon began testicular recrudescence within one month of spawning with the GSI reaching prespawning levels by September; reproductive hormones were at prespawning levels by the end of June. It seems that hydroelectric works has complex effects on sturgeon in the Mattagami system. The extensive migratory behaviour of lake sturgeon within the study area make it prone to impingement or entrainment whereas the altered river flow appears to enhance reproductive development.