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Life history plasticity in chinook salmon: relation of size and growth rate to autumnal smolting
Aquaculture 222 (2003) 149 – 165 www.elsevier.com/locate/aqua-online
Life history plasticity in chinook salmon: relation of size and growth rate to autumnal smolting Brian R. Beckman *, Donald A. Larsen, Walton W. Dickhoff Integrative Fish Biology Program, Northwest Fisheries Science Center, National Marine Fisheries Service, 2725 Montlake Blvd. East, Seattle, WA 98112 USA Received 31 October 2002; accepted 15 December 2002
Abstract An experiment was conducted to assess the relative influence of body size and growth rate on autumnal smolting of under-yearling spring chinook salmon. Fish were sorted into large (>85 mm) or small ( < 75 mm) size categories in July. Subsequently, some fish from each size group were reared at reduced ration, resulting in four treatments: Large-HiFeed, Large-LoFeed, Small-HiFeed, and Small-LoFeed. Starting in July, and continuing through November, length and weight, and samples for gill Na+ – K+ – ATPase, plasma T4 and IGF-I were obtained at 2-week intervals. On seven occasions throughout October and November, 10 fish from each treatment group were PIT tagged and movement patterns were assessed. Autumnal smolting clearly occurred in all treatment groups, indicated by elevated gill Na+ – K+ – ATPase activities, attainment of a silvery skin coloration, and an active swimming behavior pattern. However, fish from the Large-HiFeed treatment clearly differed in a number of respects from fish from the Small-LoFeed treatment from August through September: condition factor was higher, skin was more silvery, plasma T4 and IGFI levels were higher, and gill Na+ – K+ – ATPase activities were higher. In addition, fish from the Large-HiFeed group showed greater levels of activity than fish from the Small-LoFeed group on most occasions, in October and November. These data extend knowledge of the phenomenon of autumnal smolting in chinook salmon. Further, they demonstrate that differences in growth through the summer – autumn period has a significant effect on both physiological and behavioral expression of smolting. D 2003 Published by Elsevier Science B.V. Keywords: Chinook salmon; Growth; IGF-I; Plasticity; Smolting
* Corresponding author. Tel.: +1-206-860-3461; fax: +1-206-860-3467. E-mail address: [email protected] (B.R. Beckman). 0044-8486/03/$ - see front matter D 2003 Published by Elsevier Science B.V. doi:10.1016/S0044-8486(03)00108-X
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1. Introduction The life history of anadromous salmonids is marked by transitions between fresh water and seawater. The first transition occurs as juvenile fish leave fresh water habitats to enter the ocean. This is mediated by the process of smolting, a developmental event that stimulates fish to enter seawater and allows them to hypo-osmoregulate and grow after they’ve reached the ocean. Among and within salmonid species, the age, size and season at which these transitions occur may vary (L’Abe´e-Lund et al., 1989; Metcalfe and Thorpe, 1990). Chinook salmon display an especially wide variation in smolting, with fish entering seawater at sizes ranging from 1 to 30 g and at ages ranging from 30 days to 14 months post-emergence (Healey, 1991). In general, fish from northern populations smolt at an older age and larger size than fish from southern populations. However, variation appears greatest in the center of the chinook salmon’s geographic range: Southern British Columbia, Washington and Oregon. Here, variation in age, size, and season of smolting differs both between populations and within populations, between individuals (Reimers, 1973; Carl and Healey, 1984; Taylor, 1990). Traditionally, plasticity of smolting in salmonids has been related to the attainment of a threshold size (Elson, 1957). Animals were thought incapable of smolting until this size was reached. This makes logical sense from an ecological or selective standpoint. Larger smolts have greater swimming capacity, conferring greater predator avoidance and prey capture abilities. In addition, larger fish display a greater ability to withstand the challenges of seawater entry. Thus larger smolts should be better able to survive the rigors of downstream migration and ocean entry (Bilton et al., 1982; Ward et al., 1989). However, it is difficult to couple a given body size to the endocrine and physiological mechanisms which actually mediate the process of smolting. In salmonids, some evidence suggests that relatively high growth rates accelerate smolting; whereas relatively low growth rates act to retard smolting (Komourdjian et al., 1976b; Thorpe, 1977). In contrast to size, growth is directly mediated by the endocrine system. It has been suggested that salmon may physiologically couple growth to smolting through the actions of the growth hormone (GH)/insulin-like growth factor I (IGF-I) endocrine axis (Komourdjian et al., 1976b; Dickhoff et al., 1997). GH is a peptide hormone released from the pituitary; one of the primary actions of GH is to stimulate production of another peptide hormone, IGF-I from the liver. In salmon, both GH and IGFI are related to seawater tolerance, a key characteristic of salmon smolts (Komourdjian et al., 1976a; McCormick, 1996; Bjo¨rnsson, 1997). GH and IGF-I are also the major modulators of endocrine growth in fish (McLean and Donaldson, 1993; Duan, 1997). Together, these data suggest that smolt plasticity may be related to activity of the GH/ IGF-I endocrine axis. More specifically, they suggest that smolting may be induced by environmental conditions that promote high growth rates. Based on this hypothesis, we investigated the plasticity of autumnal smolting in Quilcene Hatchery spring chinook salmon. We chose to investigate autumnal smolting as it has been previously demonstrated to be plastic (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). We size-graded 0age parr in July, yielding groups of relatively large and small fish. Subsequently, each size group was divided and fed at relatively high or low ration, yielding relatively fast and slow-growing groups of relatively large and small fish. We predicted that autumnal
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smolting would be contingent on growth rate rather than size, with faster growing fish smolting while slower growing fish would not. We report our results herein.
2. Materials and methods 2.1. Fish rearing Approximately 6000 eyed spring chinook salmon eggs were obtained from the Quilcene National Fish Hatchery (Quilcene Washington, USA) in November 1995 and transported to the NMFS research hatchery in Seattle, WA. The Quilcene hatchery stock was originated by crossing Nooksack (Puget Sound) and Cowlitz River (Lower Columbia River) spring chinook salmon. Eggs and alevins were maintained in Heath Trays until December, then fry were transferred to 1.3-m diameter circular tanks and feeding was initiated. Fish were initially reared in dechlorinated Seattle city water, with a seasonally increasing temperature from 7 to 11 jC. In April, rearing water was switched to a recirculating freshwater system and fish were maintained in this system for the duration of the experiment. Temperature ranged from 11 jC to a peak of 15 jC in July, and then subsequently declined to 9 jC in November. Fish were sorted into large (>85 mm) and
Fig. 1. Length of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, SmallHiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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Fig. 2. Condition factor (weight (g) length (mm) 3 100,000) of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
small ( < 75 mm) size categories on 2 July 1996 and stocked into four 1.3-m tanks at 440 (large) and 620 (small) fish per tank (two tanks for each size category). Fish were fed BioDiet1 starter and grower diets (Bioproducts, Warrenton, OR) at the manufacturer’s recommended levels through 25 July. Subsequently, one tank from each size group was placed on a reduced ration, these low ration fish were fed the same amount per day as high-ration fish (2.2% body weight/day) but only 3 days/week (as opposed to 5 days/ week). This resulted in four experimental groups: Large-HiFeed, Large-LoFeed, SmallHiFeed, and Small-LoFeed. 2.2. Physiological assessment Beginning on 25 July, and following at approximately 2-week intervals, 12 fish were randomly dip-netted from each tank, placed in a holding bucket, and then placed singly into a lethal concentration (0.02%) of tricaine methanesulfonate (MS-222). Fish were weighed, measured, and visually assessed for smolt development (1 = parr, 2 = transitional, 3 = smolt). The tail was cut, blood was collected in heparinized glass tubes and centrifuged 1
Reference to trade name does not imply endorsement by NOAA, NMFS.
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for 3 min at 3000 g, plasma was removed and stored frozen at 80 jC. Gill tissue was cut from gill arches, placed in a buffer solution (Schrock et al., 1994), frozen and stored at 80 jC. Total unextracted plasma IGF-I concentration was determined according to Moriyama et al. (1994). Plasma T4 concentrations were determined according to Dickhoff et al. (1982). Gill Na+ – K+ – ATPase activities were measured according to Schrock et al. (1994). A laboratory accident resulted in loss of samples from two dates. 2.3. Behavioral assessment A 20-m by 1.3-m outdoor concrete raceway was modified to assess juvenile migrational behavior. A plywood barrier was positioned longitudinally down the center of the raceway. Half of a 1.3-m fibreglass tank was placed in each end to provide rounded corners. Water depth was approximately 30 cm through the center section. In each end were 60-cm deep kettles; these were approximately 2.5 m long. One pump was placed in each end to provide water flow (500 l per min). The outflowing current was routed through an array of 2 cm diameter by 20 cm length plastic pipe to straighten the flow. Water flowed from the flowstraightener longitudinally down the raceway to the inlet siphon of the second pump. Back-eddies also occurred in each corner, with water flowing directly from the flowstraightener to the inlet siphon of the same pump. The pump intake was located inside a
Fig. 3. Skin silvering index of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, LargeLoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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screened plywood box located at the bottom of the kettle. A single PIT-tag reader was placed on one side of the raceway. Fish were constrained to enter the reader through a 15 cm diameter by 50 cm length white plastic pipe. The height of the reader was adjusted so that water filled 66% of the entry pipe. Fish were PIT tagged 36 or 108 h prior to being placed in the raceway. There was no mortality associated with tagging. Passage of PITtagged fish through the tag reader was monitored for 48 –72 h, after which the raceway was drained and all fish were removed. Water temperature ranged from 13 to 16.5 jC, depending on air temperature and weather conditions. Trials occurred in October and November, when cooling air temperatures allowed water in the raceway to remain within the tolerance limits for chinook salmon. 2.4. Statistical analysis All results were first analyzed by a three-way ANOVA, with date, size, and ration the main effects. If significant effects were found ( P < 0.05), individual means were examined by a one-way ANOVA for significant differences between treatments on a given date, or significant differences between dates for a given treatment. Condition factor was calculated as weight (g) length (mm) 3 100,000. PIT-tag readings for individual fish were edited. Individual fish would take up residence in the PIT-tag reader tunnel at night.
Fig. 4. Plasma thyroxine (T4) concentration of four treatment groups of Quilcene spring chinook salmon (LargeHiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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This would generate a large number of counts within a very short period. Any series of counts for an individual fish, obtained at intervals less than 40 s, were deleted from the data. Average lap counts were log transformed prior to analysis by ANOVA. The relative overall frequency of high or low lap counts for individual fish within a treatment, for a given trial, was assessed by forming a contingency table categorizing total individual counts as active (>1 SD above the overall mean count of a trial), inactive (>1 SD below the overall mean count of a trial), or intermediate (within 1 SD of the mean). Differences between treatments in frequency of distribution between these categories were assessed by chi-square. The relations between lap count and length were assessed by simple regression. All statistical analysis were conducted with Statview 4.5 (SAS Institute). All data are expressed as means ( F standard error).
3. Results 3.1. Physiological assessment The size grading resulted in two distinct size groups of fish, as found on the first sampling in July (Fig. 1). Large and small fish remained distinctly different through
Fig. 5. Plasma insulin-like growth factor-I (IGF-I) concentration of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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samples taken in August. Beginning in September, mean sizes of large and small groups, fed different rations began to diverge. Mean lengths of fish sampled from the four groups were significantly different from each other in September and the first part of October. By October and November, mean size of Large-LoFeed and Small-HiFeed fish were not significantly different, while mean length of fish from the Large-HiFeed and SmallLoFeed groups remained distinct. Condition factors for fish from all four groups were similar at the first sample (Fig. 2). On subsequent sampling dates, fish from LoFeed groups generally had lower condition factors than fish from HiFeed groups, though differences were not always significant. There were no significant differences between mean treatment values for the final three samples taken in October and November. There were no differences in the mean skin silvering index for fish sampled on the first two dates (Fig. 3). Fish from the Small-LoFeed treatment were generally less silvered than fish from the other three groups for the remainder of the experiment. Large fish achieved the maximum silvering index of 3 in late October and November. Fish from the SmallHiFeed group also reached peak silvering index values in late October and November. Plasma T4 values were related to fish size rather than feeding level (Fig. 4). Values for the two large fish groups were similar, and significantly higher than values found in the two small fish groups for the first two sampling dates. In general, values for the large fish
Fig. 6. Gill Na+ – K+ – ATPase activity of four treatment groups of Quilcene spring chinook salmon (LargeHiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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groups fluctuated around 5 ng/ml for the remainder of the sampling. In contrast, values found in the small fish tended to fluctuate around a value of 3.5 ng/ml. There were no clear seasonal trends in any one group. Plasma IGF-I values were significantly affected by feeding level (Fig. 5). Plasma IGF-I values in Small-HiFeed fish increased significantly from levels found in Small-LoFeed fish by late August and remained significantly higher through October. Plasma IGF-I values for Large-HiFeed and Large-LoFeed fish were similar in August and September. Subsequently, plasma IGF-I values in the Large-LoFeed fish were significantly less than that of the Large-HiFeed fish in October and November. Values found in the SmallHiFeed and Large-LoFeed groups were generally similar, from late September through November. Gill Na+ – K+ – ATPase activities of large fish and small fish differed at the first sampling ( F = 4.8, p = 0.03), prior to the initiation of feeding treatments. Mean gill Na+ – K+ – ATPase values of fish from the Small-LoFeed group were significantly less than mean values found in fish from the other groups in September (Fig. 6). Mean gill Na+ – K+ – ATPase value from fish from all groups showed peak values in late October and November. Mean values for all treatments showed a significant decline from the early to the late November samples.
Fig. 7. Hourly activity of PIT-tagged salmon smolts in a modified raceway, measured on seven occasions in October and November.
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Fig. 8. Average individual activity of four treatment groups of PIT-tagged Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed) measured on seven occasions in October and November. For each trial, bars with different letters are significantly different (activity measures were log transformed prior to analysis to reduce skewness, one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
3.2. Behavioral assessment Fish movement tended to be diurnal, with peak activity during the dawn and dusk period (Fig. 7). Movement patterns tended to decrease with time spent in the raceway, with little activity after 3 days. Fish tended to move in coherent groups (schools), actively Table 1 Relation between fish size and lap count Trial
P
r2
1 2 3 4 5 6 7
0.006 0.001 0.24 0.001 0.03 0.81 0.85
0.17 0.26 – 0.43 0.10 – –
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swimming down the length of the raceway. Schools tended to break-up and reform when they reached the turbulence of a corner, some fish continuing while others stayed in the deeper water of the corner. Significant differences in movement pattern were found between trials, feeding rate, and size (three-way ANOVA trial, P < 0.0001 F = 39.1; feeding rate, P < 0.0001 F = 34.3; size, P = 0.03 F = 4.9). Significant differences were found between treatment groups for five of seven trials (Fig. 8). In each case, the BigHiFeed group had the highest average lap count. On two occasions, the Big-LoFeed treatment group had the lowest count; while on the three other occasions, the SmallLoFeed group had the lowest average count. Regression of fish length versus lap count revealed a significant relation for four of seven trials (Table 1), with the regression coefficient ranging from 0.10 to 0.43.
Fig. 9. Number of fish within a treatment group (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed) assessed as either active (>1 standard deviation above the average activity for a given trial) or inactive ( < 1 standard deviation below the average activity for a given trial).
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The frequency with which individual fish from different treatments were found in high or low activity categories is shown in Fig. 9. For every trial, fish from the Large-HiFeed treatment were found in the active category (greater than 1 SD above the mean lap count for a given trial). In no case was a fish from this treatment found in the inactive category (more than 1 SD below the mean lap count for a given trial). In contrast, fish from the other three treatments were found in all three categories. In particular, fish from the SmallHiFeed group were distributed into both active and inactive categories on six of seven trials. Similarly, fish from the Small-LoFeed groups were distributed in both categories for five of seven trials. Overall, the distribution of fish from different treatments into the different count categories was uneven (v2 = 31.5, p < 0.001).
4. Discussion Autumnal smolting clearly occurred in under-yearling Quilcene spring chinook salmon. This was indicated by elevated gill Na+ –K+ –ATPase activities, attainment of a silvery skin coloration, and an active behavior pattern. This is not a novel result; autumnal smolting has previously been demonstrated in spring chinook salmon (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). However, the demonstration of autumnal smolting was not our goal; we wished to discern whether differences in size or growth rate altered the expression of autumnal smolting. Grading fish in July clearly resulted in two different size groups of fish. Furthermore, feeding these fish at relatively high and low levels resulted in distinct groups of large and small fish with relatively high and low growth trajectories, respectively. HiFeed fish had higher plasma IGF-I levels than LoFeed fish from late September through November. Moreover, during this period, fish from the HiFeed treatments showed more active behavior patterns than fish from the LoFeed treatments. Other indicators of smolting (gill Na+ – K+ – ATPase, T4, and condition factor) showed smaller treatment differences. Thus, we did not find distinct differences in smolting between treatment groups. In several respects, these results are similar to those found for spring smolting yearling spring chinook salmon (Beckman et al., 1998a,b). Using a similar experimental design, they found that faster growing chinook salmon expressed higher plasma IGF-I levels than slower growing fish. Moreover, faster growing fish displayed a greater propensity for downstream migration than slower growing fish. Thus, both sets of experiments suggest faster growing fish, with higher plasma IGF-I levels, display greater levels of activity. Fish in the raceway exhibited activity patterns similar to fish migrating naturally in streams or rivers. They schooled: fish moved in tight directed groups, either around the raceway in circles or back and forth between the two ends, on just one side of the raceway. Records of passage through the PIT-tag detector were obviously clumped, with up to 40 fish moving through the reader within a period of 20 s. After the passage of a school, several minutes might pass before another fish passed through the reader. At any one time, up to three distinct schools could be observed. These fish behaved similarly to migrating smolts we observed in a natural stream (Beckman et al., 1998a). Moreover, directed fish movement showed daily cycles, with movement associated with daily changes in light levels (Fig. 7). The link of chinook salmon smolt passage across dams or weirs to daily photoperiod
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change has been noted previously (Independent Science Group, 1996). Based on these observations, we suggest the activity patterns we measured, which may not be strictly analogous to downstream migratory behavior, are nonetheless indicative of smolting. Beckman et al. (1998b), similar to this study, found little difference between size and growth treatments in many physiological indicators of smolting: gill Na+ – K+ – ATPase, thyroid hormone, condition factor, liver glycogen, or body lipid. Previous studies with spring chinook salmon juveniles, conducted in the autumn (with under-yearling fish) and spring (with yearling fish) have demonstrated differences in a suite of smolt characters (including gill Na+ –K+ – ATPase) associated with size or growth rate (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). The work presented here was with a group of chinook salmon with an undescribed smolting pattern. Thus, it was difficult to predict whether a given size/growth regime would or would not result in smolting. Nevertheless, we continued our investigation of the phenomena of autumnal smolting, which, in contrast to spring smolting, is relatively poorly understood. In particular, the temporal boundaries for the autumnal ‘‘smolt window’’ are as yet, unclear, making it difficult to determine when to initiate and terminate experimental treatments. Gill Na+ –K+ –ATPase values in large fish were significantly greater than small fish on the first sampling in late July, prior to manipulation of growth rates. However, given that the large fish were indeed larger than the small fish, differences in growth rate had occurred in the months between emergence and sorting in July. These gill Na+ –K+ – ATPase differences suggest that smolting was already initiated when we started our experiment. This has several implications. First, it suggests that large fish may smolt earlier than small fish, which in this case, is the same as saying that faster growing fish smolt sooner than slower growing fish. This, rather indirectly, validates our thesis: that fall smolting is conditional upon growth rate. Second, it directly suggests our experiment was flawed. We planned to alter growth rates prior to the initiation of smolting and then subsequently assess smolting differences between treatment groups. Instead, what we did was split the summer –fall period into two growth stanzas early (before July) and late (after July) and examine the influence of differential growth rates during these two periods on smolting. Thus, we actually examined the influence of growth differences on the progression of smolting, not the initiation of smolting. Fish from the Large-HiFeed treatment clearly differed in a number of respects from fish from the Small-LoFeed treatment from August through September; condition factor was higher, skin was more silvery, plasma T4 and IGF-I levels were higher, and gill Na+ –K+ – ATPase activities were higher. In October and November, condition factor became similar, skin silvering increased in both groups (though the silvering index in Small-LoFeed fish was still significantly less than that of the Large-HiFeed fish), T4 levels increased in both groups (though significant differences remained between groups), differences in plasma IGF-I levels were maintained, and gill Na+ –K+ –ATPase activities showed similar patterns of increase and decline. Finally, fish from the Large-HiFeed group showed greater levels of activity than that of the Small-LoFeed group on most occasions, October through November. Overall, it is not difficult to discern that the two groups were different: fish from the Large-HiFeed group displayed physiological traits consistent with smolting over a longer period than did the fish from the Small-LoFeed group. Moreover, fish from the Large-HiFeed treatment showed a more consistent display of behavior associated with
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smolting than did fish from the Small-LoFeed group. Thus, differences in growth rate through the summer – fall influenced the magnitude and duration of smolt-related behavioral and physiological traits. Nonetheless, fish from the Small-LoFeed group did smolt: indicated by the increase in gill Na+ – K+ – ATPase and skin silvering in October. This suggests that at least some of the fish in the Small-LoFeed group were smolts in October. However, the data on activity pattern suggests some fish were not smolts. Fish from the Large-HiFeed group show a consistent pattern of behavior, some individuals for each trial show activity greater than one standard deviation from the mean activity of that trial (Fig. 9). No individual from the Large-HiFeed group showed an activity less than one standard deviation from the mean of that trial at any time. In contrast, individuals from the Large-LoFeed, Small-HiFeed, and Small-LoFeed groups were distributed both above and below one standard deviation of the mean activity of a trial on many or most occasions. This suggests that some fish from these treatments were smolts while others were not. Fundamentally, it suggests that some individuals had passed the threshold for smolting and others had not. Thus smolting is not a property of the treatment (i.e. the treatment directly causes fish to smolt); rather, smolting is conditional upon the status of the individual fish within the treatment. The effect of the treatments is to change the distribution of individual fish status (growth) within the treatment population. Plasma IGF-I levels were not indicative of smolting. During October, when gill Na+ – + K – ATPase activities were elevated in all groups, plasma IGF-I values were low in the Large-LoFeed and Small-LoFeed groups. Thus, fish from these groups appeared to have smolted without an increase in plasma levels of IGF-I. This might suggest that plasma IGF-I levels are unrelated to smolting. Similarly, there was no obvious ‘‘peak’’ in plasma T4 levels. Certainly, there is abundant evidence that T4 is involved in the smolting process (Dickhoff et al., 1982). Evidence is also accumulating that IGF-I may be directly involved in the smolting process (Sakamoto and Hirano, 1993; McCormick, 1996). The lack of obvious peaks in plasma hormone levels does not suggest that these hormones are not involved in the smolting process, it merely suggest that changes in plasma hormone level may not be valid markers for the smolting process (Beckman et al., 1999). IGF-I levels responded to feeding levels as seen previously in fish (Pe´rez-Sa´nchez et al., 1995; Beckman et al., 1998a). Moreover, while IGF-I levels in the LoFeed fish did not increase, this does not directly suggest that the IGF-I levels were low. The Small-LoFeed group maintained a stable IGF-I level throughout the experiment, these fish also grew at a steady rate. The Small-HiFeed fish showed an initial increase in IGF-I level in response to increased feeding rates. Subsequently, IGF-I levels in these fish remained stable September through November and appear related to the higher growth rate of Small-HiFeed fish in comparison to the Small-LoFeed fish. It is possible that a greater ration restriction would have resulted in lower plasma IGF-I levels and sufficient enough suppression of the GH/ IGF-I endocrine axis to restrict smolting. We intend to further investigate this hypothesis. The phenomenon of autumnal smolting is noteworthy because it provides a unique model to study environmental effects on smolting. In general, increasing photoperiod and temperature are seasonal cues that synchronize and stimulate smolting in the spring (Hoar, 1988). In contrast, temperature and photoperiod decrease during the summer –fall period. Thus, autumnal smolting occurs under exactly opposite environmental conditions that are thought
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to stimulate spring smolting. Maturing salmon respond to decreasing photoperiod in the autumn as a cue for synchronizing spawning, so salmonids have the physiological capacity to respond to decreasing photoperiods. Clarke et al. (1989) suggested that smolting of underyearling ocean-type chinook is photoperiod-independent. Clarke and Shelbourn (1985) found that these fish developed high salinity tolerance in May – June, dependent on size (growth rate). In the present study, we found evidence of gill Na+ – K+ – ATPase differences between large and small fish in late July, about 1 month past the summer solstice, prior to any large change in photoperiod. Fish from all treatment groups showed peak gill Na+ –K+ – ATPase values in late October, approximately 1 month past the fall equinox, the time of greatest daily change in photoperiod. The relatively small differences in gill Na+ –K+ – ATPase found in July are consistent with a relation between growth and smolting, but are unlikely to be related to a decreasing photoperiod. The ‘‘peak’’ gill Na+ – K+ – ATPase activities found in all groups in October could be related to changing photoperiod but we found little difference in gill Na+ – K+ – ATPase in October between groups displaying different growth rates. The October ‘‘peak’’ appears to be size/growth independent, suggesting that the peak was dependent on some environmental cue, with photoperiod being the most obvious. However, the present experiment does not resolve the issue. It is quite clear that autumnal smolting occurs in chinook salmon; however, neither the genetic, endocrine, physiological, nor environmental cues/mechanisms responsible for this phenomenon are clear. We continue to think that smolt plasticity is in some manner related to growth and the GH/IGF-I endocrine axis. However, we need to better establish the size/ growth criteria related to smolting thresholds in particular populations before we can convincingly demonstrate this. Moreover, it does not appear that plasma IGF-I levels provide a clear signal with regard to smolting, suggesting that we need to find a different way to assess the activity of the GH/IGF-I axis with regard to smolting. Overall, we remain excited about the possibilities that autumnal smolting provide: both as a model for phenotypic plasticity and it’s endocrine control and as a comparative model for spring smolting. References Beckman, B.R., Dickhoff, W.W., 1998. Plasticity of smolting in spring chinook salmon: relation to growth and insulin-like growth factor-I. J. Fish Biol. 53, 808 – 826. Beckman, B.R., Larsen, D.A., Lee-Pawlak, B., Dickhoff, W.W., 1998a. The relation of fish size and growth rate to migration of spring chinook salmon smolts. North Am. J. Fish. Manage. 18, 537 – 546. Beckman, B.R., Larsen, D.A., Lee-Pawlak, B., Moriyama, S., Dickhoff, W.W., 1998b. Insulin-like growth factorI and environmental modulation of growth during smoltification of spring chinook salmon, (Oncorhynchus tshawytscha). Gen. Comp. Endocrinol. 109, 325 – 335. Beckman, B.R., Dickhoff, W.W., Zaugg, W.S., Sharpe, C., Hirtzel, S., Schrock, R., Larsen, D.A., Ewing, R.D., Palmisano, A., Schreck, C.B., Mahnken, C.V.W., 1999. Growth, smoltification, and smolt-to-adult return of spring chinook salmon (Oncorhynchus tshawytscha) from hatcheries on the Deschutes River, Oregon. Trans. Am. Fish. Soc. 128, 1125 – 1150. Bilton, H.T., Alderdice, D.F., Schnute, J.T., 1982. Influence of time and size at release of juvenile coho salmon (Oncorhynchus kisutch) on returns at maturity. Can. J. Fish. Aquat. Sci. 39, 426 – 447. Bjo¨rnsson, B.T., 1997. The biology of salmon growth hormone: from daylight to dominance. Fish Physiol. Biochem. 17, 9 – 24. Carl, L.M., Healey, M.C., 1984. Differences in enzyme frequency and body morphology among three juvenile
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life history types of chinook salmon (Oncorhynchus tshawytscha) in the Nanaimo river, British Columbia. Can. J. Fish. Aquat. Sci. 41, 1070 – 1077. Clarke, W.C., Shelbourn, J.E., 1985. Growth and development of seawater adaptability by juvenile fall chinook salmon (Oncorhynchus tshawytscha) in relation to temperature. Aquaculture 45, 21 – 31. Clarke, W.C., Shelbourn, J.E., Ogasawara, T., Hirano, T., 1989. Effect of initial daylength on growth, seawater adaptability and plasma growth hormone levels in underyearling coho, chinook, and chum salmon. Aquaculture 82, 51 – 62. Dickhoff, W.W., Folmar, L.C., Mighell, J.L., Mahnken, C.V.W., 1982. Plasma thyroid hormones during smoltification of yearling and under-yearling coho salmon and yearling chinook salmon and steelhead trout. Aquaculture 28, 39 – 48. Dickhoff, W.W., Beckman, B.R., Larsen, D.L., Duan, C., Moriyama, S., 1997. The role of growth in endocrine regulation of salmon smoltification. Fish Physiol. Biochem. 17, 231 – 236. Duan, C., 1997. The insulin-like growth factor system and its biological actions in fish. Am. Zool. 37, 491 – 503. Elson, P.F., 1957. The importance of size in the change from parr to smolt in Atlantic salmon. Can. Fish Cult. 21, 1 – 6. Ewing, R.D., Johnson, S.L., Pribble, H.J., Lichatowich, J.A., 1979. Temperature and photoperiod effects on gill (Na+ K+) – ATPase activity in chinook salmon (Oncorhynchus tshawytscha). J. Fish. Res. Board Can. 36, 1347 – 1353. Ewing, R.D., Pribble, H.J., Johnson, S.L., Fustish, C.S., Diamond, J., Lichatowich, J.A., 1980. Influence of size, growth rate, and photoperiod on cyclic changes in gill (Na+ K+) – ATPase activity in chinook salmon (Oncorhynchus tshawytscha). Can. J. Fish. Aquat. Sci. 37, 600 – 605. Healey, M.C., 1991. Life history of chinook salmon (Oncorhynchus tshawytscha). In: Groot, C., Margolis, L. (Eds.), Pacific Salmon Life Histories. University of British Columbia Press, Vancouver, BC, pp. 311 – 394. Hoar, W.S., 1988. The physiology of smolting salmonids. In: Hoar, W.S., Randall, D.J. (Eds.), Fish Physiology. The Physiology of Developing Fish Part B, vol. XI. Academic Press, San Diego, CA, pp. 275 – 344. Independent Science Group, 1996. Return to the river: restoration of salmonid fishes in the Columbia River ecosystem. Northwest Power Planning Council, Portland, Oregon. 96-6. http://www.nwcouncil.org/library/ index.htm. Komourdjian, M.P., Saunders, R.L., Fenwick, J.C., 1976a. The effect of porcine somatotropin on growth and survival in seawater of Atlantic salmon (Salmo salar) parr. Can. J. Zool. 54, 531 – 535. Komourdjian, M.P., Saunders, R.L., Fenwick, J.C., 1976b. Evidence for the role of growth hormone as a part of a ‘light – pituitary axis’ in growth and smoltification of Atlantic salmon (Salmo salar). Can. J. Zool. 54, 544 – 551. L’Abe´e-Lund, J.H., Jonsson, B., Jensen, A.J., Saettem, L.M., Heggegerget, T.G., Johnsen, B.O., Naesje, T.F., 1989. Latitudinal variation in life history characteristics of sea-run migrant brown trout Salmo trutta. J. Anim. Ecol. 68, 1983 – 1987. McCormick, S.D., 1996. Effects of growth hormone and insulin-like growth factor I on salinity tolerance and gill Na+, K + – ATPase in Atlantic salmon (Salmo salar): interaction with cortisol. Gen. Comp. Endocrinol. 101, 3 – 11. McLean, E., Donaldson, E.M., 1993. The role of growth hormone in growth of poikilotherms. In: Schriebman, M.P., Scanes, C.G., Pang, P.K.T. (Eds.), The Endocrinology of Growth, Development and Metabolism in Vertebrates. Academic Press, New York, pp. 43 – 71. Metcalfe, N.B., Thorpe, J.E., 1990. Determinants of geographical variation in the age of seaward-migration in Atlantic salmon, Salmo salar. J. Anim. Ecol. 59, 135 – 145. Moriyama, S., Swanson, P., Nishii, M., Takahashi, A., Kawauchi, H., Dickhoff, W.W., Plisetskaya, E.M., 1994. Development of a homologous radioimmunoassay for coho salmon insulin-like growth factor-I. Gen. Comp. Endocrinol. 96, 149 – 161. Pe´rez-Sa´nchez, J., Marti-Palanca, H., Kaushik, S.J., 1995. Ration size and protein intake affect circulating growth hormone concentration, hepatic growth hormone binding and plasma insulin-like growth factor-I immunoreactivity in a marine teleost, the gilthead sea bream (Sparus aurata). J. Nutr. 125, 546 – 552. Reimers, P.E., 1973. The length of residence of juvenile fall chinook salmon in Sixes River, Oregon. Oregon Fish Commission Res. Rep. 4, 1 – 43. Sakamoto, T., Hirano, T., 1993. Expression of an insulin-like growth factor I gene in osmoregulatory organs
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during seawater adaptation of the salmonid fish: possible mode of osmoregulatory action of growth hormone. Proc. Natl. Acad. Sci. 90, 1912 – 1916. Schrock, R.M., Beeman, J.W., Rondorf, D.W., Haner, P.V., 1994. A microassay for gill sodium, potassiumactivated ATPase in juvenile Pacific salmonids. Trans. Am. Fish. Soc. 123, 223 – 229. Taylor, E.B., 1990. Environmental correlates of life history variation in juvenile chinook salmon, Oncorhynchus tshawytscha (Walbaum). J. Fish Biol. 37, 1 – 17. Thorpe, J.E., 1977. Bimodal distribution of length of juvenile Atlantic salmon (Salmo salar L.) under artificial rearing conditions. J. Fish. Biol. 11, 175 – 184. Ward, B.R., Slaney, P.A., Facchin, A.R., Land, R.W., 1989. Size-biased survival in steelhead trout (Oncorhynchus mykiss): back-calculated lengths from adults’ scales compared to migrating smolts at the Keogh River, British Columbia. Can. J. Fish. Aquat. Sci. 46, 1853 – 1858.
Life history plasticity in chinook salmon: relation of size and growth rate to autumnal smolting Brian R. Beckman *, Donald A. Larsen, Walton W. Dickhoff Integrative Fish Biology Program, Northwest Fisheries Science Center, National Marine Fisheries Service, 2725 Montlake Blvd. East, Seattle, WA 98112 USA Received 31 October 2002; accepted 15 December 2002
Abstract An experiment was conducted to assess the relative influence of body size and growth rate on autumnal smolting of under-yearling spring chinook salmon. Fish were sorted into large (>85 mm) or small ( < 75 mm) size categories in July. Subsequently, some fish from each size group were reared at reduced ration, resulting in four treatments: Large-HiFeed, Large-LoFeed, Small-HiFeed, and Small-LoFeed. Starting in July, and continuing through November, length and weight, and samples for gill Na+ – K+ – ATPase, plasma T4 and IGF-I were obtained at 2-week intervals. On seven occasions throughout October and November, 10 fish from each treatment group were PIT tagged and movement patterns were assessed. Autumnal smolting clearly occurred in all treatment groups, indicated by elevated gill Na+ – K+ – ATPase activities, attainment of a silvery skin coloration, and an active swimming behavior pattern. However, fish from the Large-HiFeed treatment clearly differed in a number of respects from fish from the Small-LoFeed treatment from August through September: condition factor was higher, skin was more silvery, plasma T4 and IGFI levels were higher, and gill Na+ – K+ – ATPase activities were higher. In addition, fish from the Large-HiFeed group showed greater levels of activity than fish from the Small-LoFeed group on most occasions, in October and November. These data extend knowledge of the phenomenon of autumnal smolting in chinook salmon. Further, they demonstrate that differences in growth through the summer – autumn period has a significant effect on both physiological and behavioral expression of smolting. D 2003 Published by Elsevier Science B.V. Keywords: Chinook salmon; Growth; IGF-I; Plasticity; Smolting
* Corresponding author. Tel.: +1-206-860-3461; fax: +1-206-860-3467. E-mail address: [email protected] (B.R. Beckman). 0044-8486/03/$ - see front matter D 2003 Published by Elsevier Science B.V. doi:10.1016/S0044-8486(03)00108-X
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1. Introduction The life history of anadromous salmonids is marked by transitions between fresh water and seawater. The first transition occurs as juvenile fish leave fresh water habitats to enter the ocean. This is mediated by the process of smolting, a developmental event that stimulates fish to enter seawater and allows them to hypo-osmoregulate and grow after they’ve reached the ocean. Among and within salmonid species, the age, size and season at which these transitions occur may vary (L’Abe´e-Lund et al., 1989; Metcalfe and Thorpe, 1990). Chinook salmon display an especially wide variation in smolting, with fish entering seawater at sizes ranging from 1 to 30 g and at ages ranging from 30 days to 14 months post-emergence (Healey, 1991). In general, fish from northern populations smolt at an older age and larger size than fish from southern populations. However, variation appears greatest in the center of the chinook salmon’s geographic range: Southern British Columbia, Washington and Oregon. Here, variation in age, size, and season of smolting differs both between populations and within populations, between individuals (Reimers, 1973; Carl and Healey, 1984; Taylor, 1990). Traditionally, plasticity of smolting in salmonids has been related to the attainment of a threshold size (Elson, 1957). Animals were thought incapable of smolting until this size was reached. This makes logical sense from an ecological or selective standpoint. Larger smolts have greater swimming capacity, conferring greater predator avoidance and prey capture abilities. In addition, larger fish display a greater ability to withstand the challenges of seawater entry. Thus larger smolts should be better able to survive the rigors of downstream migration and ocean entry (Bilton et al., 1982; Ward et al., 1989). However, it is difficult to couple a given body size to the endocrine and physiological mechanisms which actually mediate the process of smolting. In salmonids, some evidence suggests that relatively high growth rates accelerate smolting; whereas relatively low growth rates act to retard smolting (Komourdjian et al., 1976b; Thorpe, 1977). In contrast to size, growth is directly mediated by the endocrine system. It has been suggested that salmon may physiologically couple growth to smolting through the actions of the growth hormone (GH)/insulin-like growth factor I (IGF-I) endocrine axis (Komourdjian et al., 1976b; Dickhoff et al., 1997). GH is a peptide hormone released from the pituitary; one of the primary actions of GH is to stimulate production of another peptide hormone, IGF-I from the liver. In salmon, both GH and IGFI are related to seawater tolerance, a key characteristic of salmon smolts (Komourdjian et al., 1976a; McCormick, 1996; Bjo¨rnsson, 1997). GH and IGF-I are also the major modulators of endocrine growth in fish (McLean and Donaldson, 1993; Duan, 1997). Together, these data suggest that smolt plasticity may be related to activity of the GH/ IGF-I endocrine axis. More specifically, they suggest that smolting may be induced by environmental conditions that promote high growth rates. Based on this hypothesis, we investigated the plasticity of autumnal smolting in Quilcene Hatchery spring chinook salmon. We chose to investigate autumnal smolting as it has been previously demonstrated to be plastic (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). We size-graded 0age parr in July, yielding groups of relatively large and small fish. Subsequently, each size group was divided and fed at relatively high or low ration, yielding relatively fast and slow-growing groups of relatively large and small fish. We predicted that autumnal
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smolting would be contingent on growth rate rather than size, with faster growing fish smolting while slower growing fish would not. We report our results herein.
2. Materials and methods 2.1. Fish rearing Approximately 6000 eyed spring chinook salmon eggs were obtained from the Quilcene National Fish Hatchery (Quilcene Washington, USA) in November 1995 and transported to the NMFS research hatchery in Seattle, WA. The Quilcene hatchery stock was originated by crossing Nooksack (Puget Sound) and Cowlitz River (Lower Columbia River) spring chinook salmon. Eggs and alevins were maintained in Heath Trays until December, then fry were transferred to 1.3-m diameter circular tanks and feeding was initiated. Fish were initially reared in dechlorinated Seattle city water, with a seasonally increasing temperature from 7 to 11 jC. In April, rearing water was switched to a recirculating freshwater system and fish were maintained in this system for the duration of the experiment. Temperature ranged from 11 jC to a peak of 15 jC in July, and then subsequently declined to 9 jC in November. Fish were sorted into large (>85 mm) and
Fig. 1. Length of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, SmallHiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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Fig. 2. Condition factor (weight (g) length (mm) 3 100,000) of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
small ( < 75 mm) size categories on 2 July 1996 and stocked into four 1.3-m tanks at 440 (large) and 620 (small) fish per tank (two tanks for each size category). Fish were fed BioDiet1 starter and grower diets (Bioproducts, Warrenton, OR) at the manufacturer’s recommended levels through 25 July. Subsequently, one tank from each size group was placed on a reduced ration, these low ration fish were fed the same amount per day as high-ration fish (2.2% body weight/day) but only 3 days/week (as opposed to 5 days/ week). This resulted in four experimental groups: Large-HiFeed, Large-LoFeed, SmallHiFeed, and Small-LoFeed. 2.2. Physiological assessment Beginning on 25 July, and following at approximately 2-week intervals, 12 fish were randomly dip-netted from each tank, placed in a holding bucket, and then placed singly into a lethal concentration (0.02%) of tricaine methanesulfonate (MS-222). Fish were weighed, measured, and visually assessed for smolt development (1 = parr, 2 = transitional, 3 = smolt). The tail was cut, blood was collected in heparinized glass tubes and centrifuged 1
Reference to trade name does not imply endorsement by NOAA, NMFS.
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for 3 min at 3000 g, plasma was removed and stored frozen at 80 jC. Gill tissue was cut from gill arches, placed in a buffer solution (Schrock et al., 1994), frozen and stored at 80 jC. Total unextracted plasma IGF-I concentration was determined according to Moriyama et al. (1994). Plasma T4 concentrations were determined according to Dickhoff et al. (1982). Gill Na+ – K+ – ATPase activities were measured according to Schrock et al. (1994). A laboratory accident resulted in loss of samples from two dates. 2.3. Behavioral assessment A 20-m by 1.3-m outdoor concrete raceway was modified to assess juvenile migrational behavior. A plywood barrier was positioned longitudinally down the center of the raceway. Half of a 1.3-m fibreglass tank was placed in each end to provide rounded corners. Water depth was approximately 30 cm through the center section. In each end were 60-cm deep kettles; these were approximately 2.5 m long. One pump was placed in each end to provide water flow (500 l per min). The outflowing current was routed through an array of 2 cm diameter by 20 cm length plastic pipe to straighten the flow. Water flowed from the flowstraightener longitudinally down the raceway to the inlet siphon of the second pump. Back-eddies also occurred in each corner, with water flowing directly from the flowstraightener to the inlet siphon of the same pump. The pump intake was located inside a
Fig. 3. Skin silvering index of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, LargeLoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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screened plywood box located at the bottom of the kettle. A single PIT-tag reader was placed on one side of the raceway. Fish were constrained to enter the reader through a 15 cm diameter by 50 cm length white plastic pipe. The height of the reader was adjusted so that water filled 66% of the entry pipe. Fish were PIT tagged 36 or 108 h prior to being placed in the raceway. There was no mortality associated with tagging. Passage of PITtagged fish through the tag reader was monitored for 48 –72 h, after which the raceway was drained and all fish were removed. Water temperature ranged from 13 to 16.5 jC, depending on air temperature and weather conditions. Trials occurred in October and November, when cooling air temperatures allowed water in the raceway to remain within the tolerance limits for chinook salmon. 2.4. Statistical analysis All results were first analyzed by a three-way ANOVA, with date, size, and ration the main effects. If significant effects were found ( P < 0.05), individual means were examined by a one-way ANOVA for significant differences between treatments on a given date, or significant differences between dates for a given treatment. Condition factor was calculated as weight (g) length (mm) 3 100,000. PIT-tag readings for individual fish were edited. Individual fish would take up residence in the PIT-tag reader tunnel at night.
Fig. 4. Plasma thyroxine (T4) concentration of four treatment groups of Quilcene spring chinook salmon (LargeHiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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This would generate a large number of counts within a very short period. Any series of counts for an individual fish, obtained at intervals less than 40 s, were deleted from the data. Average lap counts were log transformed prior to analysis by ANOVA. The relative overall frequency of high or low lap counts for individual fish within a treatment, for a given trial, was assessed by forming a contingency table categorizing total individual counts as active (>1 SD above the overall mean count of a trial), inactive (>1 SD below the overall mean count of a trial), or intermediate (within 1 SD of the mean). Differences between treatments in frequency of distribution between these categories were assessed by chi-square. The relations between lap count and length were assessed by simple regression. All statistical analysis were conducted with Statview 4.5 (SAS Institute). All data are expressed as means ( F standard error).
3. Results 3.1. Physiological assessment The size grading resulted in two distinct size groups of fish, as found on the first sampling in July (Fig. 1). Large and small fish remained distinctly different through
Fig. 5. Plasma insulin-like growth factor-I (IGF-I) concentration of four treatment groups of Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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samples taken in August. Beginning in September, mean sizes of large and small groups, fed different rations began to diverge. Mean lengths of fish sampled from the four groups were significantly different from each other in September and the first part of October. By October and November, mean size of Large-LoFeed and Small-HiFeed fish were not significantly different, while mean length of fish from the Large-HiFeed and SmallLoFeed groups remained distinct. Condition factors for fish from all four groups were similar at the first sample (Fig. 2). On subsequent sampling dates, fish from LoFeed groups generally had lower condition factors than fish from HiFeed groups, though differences were not always significant. There were no significant differences between mean treatment values for the final three samples taken in October and November. There were no differences in the mean skin silvering index for fish sampled on the first two dates (Fig. 3). Fish from the Small-LoFeed treatment were generally less silvered than fish from the other three groups for the remainder of the experiment. Large fish achieved the maximum silvering index of 3 in late October and November. Fish from the SmallHiFeed group also reached peak silvering index values in late October and November. Plasma T4 values were related to fish size rather than feeding level (Fig. 4). Values for the two large fish groups were similar, and significantly higher than values found in the two small fish groups for the first two sampling dates. In general, values for the large fish
Fig. 6. Gill Na+ – K+ – ATPase activity of four treatment groups of Quilcene spring chinook salmon (LargeHiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed). Different letters indicate that significant differences were found between groups on a given date (one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
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groups fluctuated around 5 ng/ml for the remainder of the sampling. In contrast, values found in the small fish tended to fluctuate around a value of 3.5 ng/ml. There were no clear seasonal trends in any one group. Plasma IGF-I values were significantly affected by feeding level (Fig. 5). Plasma IGF-I values in Small-HiFeed fish increased significantly from levels found in Small-LoFeed fish by late August and remained significantly higher through October. Plasma IGF-I values for Large-HiFeed and Large-LoFeed fish were similar in August and September. Subsequently, plasma IGF-I values in the Large-LoFeed fish were significantly less than that of the Large-HiFeed fish in October and November. Values found in the SmallHiFeed and Large-LoFeed groups were generally similar, from late September through November. Gill Na+ – K+ – ATPase activities of large fish and small fish differed at the first sampling ( F = 4.8, p = 0.03), prior to the initiation of feeding treatments. Mean gill Na+ – K+ – ATPase values of fish from the Small-LoFeed group were significantly less than mean values found in fish from the other groups in September (Fig. 6). Mean gill Na+ – K+ – ATPase value from fish from all groups showed peak values in late October and November. Mean values for all treatments showed a significant decline from the early to the late November samples.
Fig. 7. Hourly activity of PIT-tagged salmon smolts in a modified raceway, measured on seven occasions in October and November.
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Fig. 8. Average individual activity of four treatment groups of PIT-tagged Quilcene spring chinook salmon (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed) measured on seven occasions in October and November. For each trial, bars with different letters are significantly different (activity measures were log transformed prior to analysis to reduce skewness, one-way ANOVA, followed by Fisher’s PLSD, P < 0.05).
3.2. Behavioral assessment Fish movement tended to be diurnal, with peak activity during the dawn and dusk period (Fig. 7). Movement patterns tended to decrease with time spent in the raceway, with little activity after 3 days. Fish tended to move in coherent groups (schools), actively Table 1 Relation between fish size and lap count Trial
P
r2
1 2 3 4 5 6 7
0.006 0.001 0.24 0.001 0.03 0.81 0.85
0.17 0.26 – 0.43 0.10 – –
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swimming down the length of the raceway. Schools tended to break-up and reform when they reached the turbulence of a corner, some fish continuing while others stayed in the deeper water of the corner. Significant differences in movement pattern were found between trials, feeding rate, and size (three-way ANOVA trial, P < 0.0001 F = 39.1; feeding rate, P < 0.0001 F = 34.3; size, P = 0.03 F = 4.9). Significant differences were found between treatment groups for five of seven trials (Fig. 8). In each case, the BigHiFeed group had the highest average lap count. On two occasions, the Big-LoFeed treatment group had the lowest count; while on the three other occasions, the SmallLoFeed group had the lowest average count. Regression of fish length versus lap count revealed a significant relation for four of seven trials (Table 1), with the regression coefficient ranging from 0.10 to 0.43.
Fig. 9. Number of fish within a treatment group (Large-HiFeed, Large-LoFeed, Small-HiFeed, Small-LoFeed) assessed as either active (>1 standard deviation above the average activity for a given trial) or inactive ( < 1 standard deviation below the average activity for a given trial).
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The frequency with which individual fish from different treatments were found in high or low activity categories is shown in Fig. 9. For every trial, fish from the Large-HiFeed treatment were found in the active category (greater than 1 SD above the mean lap count for a given trial). In no case was a fish from this treatment found in the inactive category (more than 1 SD below the mean lap count for a given trial). In contrast, fish from the other three treatments were found in all three categories. In particular, fish from the SmallHiFeed group were distributed into both active and inactive categories on six of seven trials. Similarly, fish from the Small-LoFeed groups were distributed in both categories for five of seven trials. Overall, the distribution of fish from different treatments into the different count categories was uneven (v2 = 31.5, p < 0.001).
4. Discussion Autumnal smolting clearly occurred in under-yearling Quilcene spring chinook salmon. This was indicated by elevated gill Na+ –K+ –ATPase activities, attainment of a silvery skin coloration, and an active behavior pattern. This is not a novel result; autumnal smolting has previously been demonstrated in spring chinook salmon (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). However, the demonstration of autumnal smolting was not our goal; we wished to discern whether differences in size or growth rate altered the expression of autumnal smolting. Grading fish in July clearly resulted in two different size groups of fish. Furthermore, feeding these fish at relatively high and low levels resulted in distinct groups of large and small fish with relatively high and low growth trajectories, respectively. HiFeed fish had higher plasma IGF-I levels than LoFeed fish from late September through November. Moreover, during this period, fish from the HiFeed treatments showed more active behavior patterns than fish from the LoFeed treatments. Other indicators of smolting (gill Na+ – K+ – ATPase, T4, and condition factor) showed smaller treatment differences. Thus, we did not find distinct differences in smolting between treatment groups. In several respects, these results are similar to those found for spring smolting yearling spring chinook salmon (Beckman et al., 1998a,b). Using a similar experimental design, they found that faster growing chinook salmon expressed higher plasma IGF-I levels than slower growing fish. Moreover, faster growing fish displayed a greater propensity for downstream migration than slower growing fish. Thus, both sets of experiments suggest faster growing fish, with higher plasma IGF-I levels, display greater levels of activity. Fish in the raceway exhibited activity patterns similar to fish migrating naturally in streams or rivers. They schooled: fish moved in tight directed groups, either around the raceway in circles or back and forth between the two ends, on just one side of the raceway. Records of passage through the PIT-tag detector were obviously clumped, with up to 40 fish moving through the reader within a period of 20 s. After the passage of a school, several minutes might pass before another fish passed through the reader. At any one time, up to three distinct schools could be observed. These fish behaved similarly to migrating smolts we observed in a natural stream (Beckman et al., 1998a). Moreover, directed fish movement showed daily cycles, with movement associated with daily changes in light levels (Fig. 7). The link of chinook salmon smolt passage across dams or weirs to daily photoperiod
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change has been noted previously (Independent Science Group, 1996). Based on these observations, we suggest the activity patterns we measured, which may not be strictly analogous to downstream migratory behavior, are nonetheless indicative of smolting. Beckman et al. (1998b), similar to this study, found little difference between size and growth treatments in many physiological indicators of smolting: gill Na+ – K+ – ATPase, thyroid hormone, condition factor, liver glycogen, or body lipid. Previous studies with spring chinook salmon juveniles, conducted in the autumn (with under-yearling fish) and spring (with yearling fish) have demonstrated differences in a suite of smolt characters (including gill Na+ –K+ – ATPase) associated with size or growth rate (Ewing et al., 1979, 1980; Beckman and Dickhoff, 1998). The work presented here was with a group of chinook salmon with an undescribed smolting pattern. Thus, it was difficult to predict whether a given size/growth regime would or would not result in smolting. Nevertheless, we continued our investigation of the phenomena of autumnal smolting, which, in contrast to spring smolting, is relatively poorly understood. In particular, the temporal boundaries for the autumnal ‘‘smolt window’’ are as yet, unclear, making it difficult to determine when to initiate and terminate experimental treatments. Gill Na+ –K+ –ATPase values in large fish were significantly greater than small fish on the first sampling in late July, prior to manipulation of growth rates. However, given that the large fish were indeed larger than the small fish, differences in growth rate had occurred in the months between emergence and sorting in July. These gill Na+ –K+ – ATPase differences suggest that smolting was already initiated when we started our experiment. This has several implications. First, it suggests that large fish may smolt earlier than small fish, which in this case, is the same as saying that faster growing fish smolt sooner than slower growing fish. This, rather indirectly, validates our thesis: that fall smolting is conditional upon growth rate. Second, it directly suggests our experiment was flawed. We planned to alter growth rates prior to the initiation of smolting and then subsequently assess smolting differences between treatment groups. Instead, what we did was split the summer –fall period into two growth stanzas early (before July) and late (after July) and examine the influence of differential growth rates during these two periods on smolting. Thus, we actually examined the influence of growth differences on the progression of smolting, not the initiation of smolting. Fish from the Large-HiFeed treatment clearly differed in a number of respects from fish from the Small-LoFeed treatment from August through September; condition factor was higher, skin was more silvery, plasma T4 and IGF-I levels were higher, and gill Na+ –K+ – ATPase activities were higher. In October and November, condition factor became similar, skin silvering increased in both groups (though the silvering index in Small-LoFeed fish was still significantly less than that of the Large-HiFeed fish), T4 levels increased in both groups (though significant differences remained between groups), differences in plasma IGF-I levels were maintained, and gill Na+ –K+ –ATPase activities showed similar patterns of increase and decline. Finally, fish from the Large-HiFeed group showed greater levels of activity than that of the Small-LoFeed group on most occasions, October through November. Overall, it is not difficult to discern that the two groups were different: fish from the Large-HiFeed group displayed physiological traits consistent with smolting over a longer period than did the fish from the Small-LoFeed group. Moreover, fish from the Large-HiFeed treatment showed a more consistent display of behavior associated with
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smolting than did fish from the Small-LoFeed group. Thus, differences in growth rate through the summer – fall influenced the magnitude and duration of smolt-related behavioral and physiological traits. Nonetheless, fish from the Small-LoFeed group did smolt: indicated by the increase in gill Na+ – K+ – ATPase and skin silvering in October. This suggests that at least some of the fish in the Small-LoFeed group were smolts in October. However, the data on activity pattern suggests some fish were not smolts. Fish from the Large-HiFeed group show a consistent pattern of behavior, some individuals for each trial show activity greater than one standard deviation from the mean activity of that trial (Fig. 9). No individual from the Large-HiFeed group showed an activity less than one standard deviation from the mean of that trial at any time. In contrast, individuals from the Large-LoFeed, Small-HiFeed, and Small-LoFeed groups were distributed both above and below one standard deviation of the mean activity of a trial on many or most occasions. This suggests that some fish from these treatments were smolts while others were not. Fundamentally, it suggests that some individuals had passed the threshold for smolting and others had not. Thus smolting is not a property of the treatment (i.e. the treatment directly causes fish to smolt); rather, smolting is conditional upon the status of the individual fish within the treatment. The effect of the treatments is to change the distribution of individual fish status (growth) within the treatment population. Plasma IGF-I levels were not indicative of smolting. During October, when gill Na+ – + K – ATPase activities were elevated in all groups, plasma IGF-I values were low in the Large-LoFeed and Small-LoFeed groups. Thus, fish from these groups appeared to have smolted without an increase in plasma levels of IGF-I. This might suggest that plasma IGF-I levels are unrelated to smolting. Similarly, there was no obvious ‘‘peak’’ in plasma T4 levels. Certainly, there is abundant evidence that T4 is involved in the smolting process (Dickhoff et al., 1982). Evidence is also accumulating that IGF-I may be directly involved in the smolting process (Sakamoto and Hirano, 1993; McCormick, 1996). The lack of obvious peaks in plasma hormone levels does not suggest that these hormones are not involved in the smolting process, it merely suggest that changes in plasma hormone level may not be valid markers for the smolting process (Beckman et al., 1999). IGF-I levels responded to feeding levels as seen previously in fish (Pe´rez-Sa´nchez et al., 1995; Beckman et al., 1998a). Moreover, while IGF-I levels in the LoFeed fish did not increase, this does not directly suggest that the IGF-I levels were low. The Small-LoFeed group maintained a stable IGF-I level throughout the experiment, these fish also grew at a steady rate. The Small-HiFeed fish showed an initial increase in IGF-I level in response to increased feeding rates. Subsequently, IGF-I levels in these fish remained stable September through November and appear related to the higher growth rate of Small-HiFeed fish in comparison to the Small-LoFeed fish. It is possible that a greater ration restriction would have resulted in lower plasma IGF-I levels and sufficient enough suppression of the GH/ IGF-I endocrine axis to restrict smolting. We intend to further investigate this hypothesis. The phenomenon of autumnal smolting is noteworthy because it provides a unique model to study environmental effects on smolting. In general, increasing photoperiod and temperature are seasonal cues that synchronize and stimulate smolting in the spring (Hoar, 1988). In contrast, temperature and photoperiod decrease during the summer –fall period. Thus, autumnal smolting occurs under exactly opposite environmental conditions that are thought
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to stimulate spring smolting. Maturing salmon respond to decreasing photoperiod in the autumn as a cue for synchronizing spawning, so salmonids have the physiological capacity to respond to decreasing photoperiods. Clarke et al. (1989) suggested that smolting of underyearling ocean-type chinook is photoperiod-independent. Clarke and Shelbourn (1985) found that these fish developed high salinity tolerance in May – June, dependent on size (growth rate). In the present study, we found evidence of gill Na+ – K+ – ATPase differences between large and small fish in late July, about 1 month past the summer solstice, prior to any large change in photoperiod. Fish from all treatment groups showed peak gill Na+ –K+ – ATPase values in late October, approximately 1 month past the fall equinox, the time of greatest daily change in photoperiod. The relatively small differences in gill Na+ –K+ – ATPase found in July are consistent with a relation between growth and smolting, but are unlikely to be related to a decreasing photoperiod. The ‘‘peak’’ gill Na+ – K+ – ATPase activities found in all groups in October could be related to changing photoperiod but we found little difference in gill Na+ – K+ – ATPase in October between groups displaying different growth rates. The October ‘‘peak’’ appears to be size/growth independent, suggesting that the peak was dependent on some environmental cue, with photoperiod being the most obvious. However, the present experiment does not resolve the issue. It is quite clear that autumnal smolting occurs in chinook salmon; however, neither the genetic, endocrine, physiological, nor environmental cues/mechanisms responsible for this phenomenon are clear. We continue to think that smolt plasticity is in some manner related to growth and the GH/IGF-I endocrine axis. However, we need to better establish the size/ growth criteria related to smolting thresholds in particular populations before we can convincingly demonstrate this. Moreover, it does not appear that plasma IGF-I levels provide a clear signal with regard to smolting, suggesting that we need to find a different way to assess the activity of the GH/IGF-I axis with regard to smolting. Overall, we remain excited about the possibilities that autumnal smolting provide: both as a model for phenotypic plasticity and it’s endocrine control and as a comparative model for spring smolting. References Beckman, B.R., Dickhoff, W.W., 1998. Plasticity of smolting in spring chinook salmon: relation to growth and insulin-like growth factor-I. J. Fish Biol. 53, 808 – 826. Beckman, B.R., Larsen, D.A., Lee-Pawlak, B., Dickhoff, W.W., 1998a. The relation of fish size and growth rate to migration of spring chinook salmon smolts. North Am. J. Fish. Manage. 18, 537 – 546. Beckman, B.R., Larsen, D.A., Lee-Pawlak, B., Moriyama, S., Dickhoff, W.W., 1998b. Insulin-like growth factorI and environmental modulation of growth during smoltification of spring chinook salmon, (Oncorhynchus tshawytscha). Gen. Comp. Endocrinol. 109, 325 – 335. Beckman, B.R., Dickhoff, W.W., Zaugg, W.S., Sharpe, C., Hirtzel, S., Schrock, R., Larsen, D.A., Ewing, R.D., Palmisano, A., Schreck, C.B., Mahnken, C.V.W., 1999. Growth, smoltification, and smolt-to-adult return of spring chinook salmon (Oncorhynchus tshawytscha) from hatcheries on the Deschutes River, Oregon. Trans. Am. Fish. Soc. 128, 1125 – 1150. Bilton, H.T., Alderdice, D.F., Schnute, J.T., 1982. Influence of time and size at release of juvenile coho salmon (Oncorhynchus kisutch) on returns at maturity. Can. J. Fish. Aquat. Sci. 39, 426 – 447. Bjo¨rnsson, B.T., 1997. The biology of salmon growth hormone: from daylight to dominance. Fish Physiol. Biochem. 17, 9 – 24. Carl, L.M., Healey, M.C., 1984. Differences in enzyme frequency and body morphology among three juvenile
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