Improved rotatory-flow technique applied to Cod (Gadus morrhua L.)

|4'ater Research Vol 10. pp 833 to 845. Pergamon Pres~ 1'~6 Printed in Great Britain

I M P R O V E D ROTATORY-FLOW TECHNIQUE APPLIED TO COD (GADUS MORRHUA L.) P. E. LINDAHL, S. OLOFSSON and E. SCHWANBOM Institute of Zoophysiology. Uppsala, Sweden

[Receired 9 July 1975~ Abstract--An improved version of rotatory-flow apparatus is described, allowing tests of fitness with fish of different size at constant temperature. Cod (Gadus morrhua, L.t was used as experimental animal. and its treatment and behaviour during different phases of the test are described in some detail. Each test results in determination of the "critical rev rain- ~'' at which the fish is just brought to rotate with the water. The mean of a series of 10 critical rev min-~ with the same fish, multiplied by the interior circumference of the rotational tube used, gives the "'critical peripheral velocity" of the specimen. Critical peripheral velocities of a sample of fish, plotted against the standard length of the fish. arrange themselves along a straight line which intersects the vertical axis near the origin. Divided by the corresponding standard lengths the critical peripheral velocities give "reaction quotients", the mean of which describe the reaction of the whole sample of fish. Optimal experimental conditions have been found by investigating the effects of systematic variation in streaming velocity, relationship between rotational tube diameter and fin-breadth, temperature, and effects of repetition of test-series on successive days etc, on the results of testing.

INTRODUCTION

at this department. The present stand is made out of steel and acryl plastics and may be taken apart in order to facilitate transport. One of the two bearings supporting the rotating tube may be moved between two positions in order to make possible the use of tubes of different lengths. The rotating tubes are supported by double bearings in order to improve stability at high rotational velocities. To be able to study specimens of varying size it was necessary to have access to tubes with different dimensions. The tubes used so far had interior diameters of 20, 40. 50, 55, 60, 65, 70, 80 and 90 mm with a wall thickness of 5 ram; the 4 narrowest tubes were 310 and the wider ones 430 mm long. Furthermore the two tubes with interior diameters of 80 and 90 mm have been provided with interior, loose, thin-walled (3 mm) tubes with the same exterior diameters and split open along the side in order to facilitate their pushing in and removal. This arrangement allows the two tubes to be used both with their original interior diameter and also with one which is 6 mm shorter. The different tubes, being mounted as previously in equally sized supporting circular discs (Fig. 1) may easily be exchanged. A stronger driving motor and a completely mechanic controlling unit make possible higher maximal rev min-1 and greater option in varying acceleration. In order to facilitate the determination of critical rev mina tachometer was mounted, showing the rev min-1 of the tubes. This could be read with an accuracy of l°o. The peripheral velocity of the interior surface of the tube is then calculated. Each rotational tube was provided with a fine-meshed net of stainless steel (size of mesh 1 mink about 1 cm behind the end of the inflow cone. In order to further diminish differences in flow rate within the transverse section of the rotational tubes and guarantee that the water mass entering the main compartment of the rotational tube adopts the rotational velocity of the latter, thinwalled (0.6 mml elastic acetylcellulose tubes of l0 mm diameter (Lusteroid centrifuge tubes from International Equipment CP, bottoms removedl were fitted into the rotational tube, perpendicularly to the net and filling out the whole transverse section (Fig. 1 i, Their number was chosen so that they were somewhat deformed, the strain thus developed keeping them in position. In the 4 narrower rotational tubes their length was 35 mm, in the remaining ones 50 mm.

L a b o r a t o r y experiments on roach demonstrated that sublethal methyl-mercury poisoning may be revealed and quantitatively estimated with the aid of a rotatoryflow technique (Lindahl and Schwanbom, 1971al. This technique implies that the fish to be studied is placed in water, streaming through a narrow horizontal tube and revolving a r o u n d its direction of flow with linearly accelerating velocity. The critical rev m i n - 1 at which the fish is just b r o u g h t to rotate with the water is read a n d used as a measure of the ability of the fish to resist the forces of torque working u p o n it. Preliminary experiments with roach indicated that the critical rev r a i n - I was negatively correlated to the concentration of C H a H g ÷ in the body muscles, A similar series of experiments performed in the field u n d e r more optimal conditions confirmed this result (Lindahl a n d Schwanbom, 1971b). The principle, a l t h o u g h with use of a modified apparatus, was also applied to the effect of zinc on minnows (Bengtsson. 1975). The present study aims at a further development of this technique and an analysis of the experimental conditions for comparison of different populations in the sea, especially of cod. This d e m a n d s close knowledge of b o t h the effect of individual properties, e.g. size, on the test reaction a n d of the importance of more general biological factors, e.g. temperature.

EXPERIMENTAL

Apparatus The apparatus described earlier (Lindahl and Schwanborn, 1971al has been substituted by a model (Fig. 1 I, improved in many respects and. as the prototype, constructed 833

834

P. E, LINDAHL, S. OLOFSSON and E. SCHWANBOM

Fig. 1. Improved model of the rotatory-flow apparatus. The water flows from the left to the right. (1) Rotating tube: (2) thin-walled inset-tubes, 10mm in diameter, filling up the whole cross section of the rotating tube; (3) ring supporting a fine-meshed net which may be pushed into any position to keep the fish near the inflow end of the rotational tube: (4) two circular discs which run in a kind of driving bearings and support the rotating tube; (5) marks the position of the right bearing when short rotating tubes are used: (6) inflow cone: (7) outflow cone: (8) tachometer generator with driving belt around the rotational tube: (9) drive unit. When once brought into rotation the current water mass mainly preserves its rotational rate. As soon as the water has passed the inset-tubes its rotational rate in each section of the rotating tube is lower than that of the corresponding section of the tube. since the former continues at about the same rate. once adopted, while the rate of the tube steadily increases. The difference at each moment between the rotational rates of tube and water at the outflow end of the rotating tube increases with increasing acceleration and length of the tube and decreases with increasing current velocity, e.g. for the long rotating tubes (430 m m 50 mm of inset-tubes), the linear water velocity 3.0 cm s- 1 and the acceleration 70 rev rain -z this difference is 14.8 rev m i n - t . In order to reduce this discrepancy between the observed rev m i n - I of the tube and that of the current of water influencing the fish. the latter should be forced to stay in the immediate vicinity of the insert-tubes. Therefore, a circular metal net mounted in a frame was provided for each tube (Fig. l). This net is introduced into the tube after the fish and is pushed in so far that the fish occupies a section of the tube exceeding the total length of the fish by about 3 cm. Here the frame is fixed by springpressure against the interior surface of the tube. In all experiments described the tubes were provided with these nets unless otherwise stated. Since the head of the fish points against the direction of flow we speak of left and right in the tube in relation to the normal position of the fish. This is also done when the fish is in other positions (p. 835). The direction of rotation of the tube is clockwise when facing the current. Experiments with different constant temperatures were made possible by using a closed circulatory system. From a storage tank, at a lower level than that of the rotating tube, the sea water was pumped up into a higher seated tank (overflow level-regulator) from where it flows back to the storage tank. partly via the tube and partly as surplus water via a bypass. In the storage tank the water is well aerated by a circulation pump. Temperature is kept

constant (_+0.5:) and independent of that of the environment by balancing the effect of an immersion heater against that of a continuous-flow refrigerator. When exchanging rotating tubes the pressure head was kept constant (about 700 mm H:O) and the water velocity regulated by changing the diameter and length of glass tubes inserted in the stopper closing the outflow cone. The linear water velocity desired was converted into volume per unit time and was then adjusted with the aid of large graduated measuring cylinders.

Experimental animals and their treatment Several species of fish. cod (Gadus morrhua, L.), fivebearded rockling (Onos mustelus, L.), viviparous blenny (Zoarces viriparus, L.), Greenland bullhead (Acanthocottus scorpius, L.), goldsinny (Stenolabrus rupestris L.) and stickleback (Gasterosteus aculeatus, L.) were examined in the rotating tube. One of the principle conditions for suitability of different species as experimental fish in the present method is a long head in relation to the rest of the body, preventing the fish from turning its head in the direction of flow. The viviparous blenny and the goldsinny did not fulfil this condition, Furthermore, only species swimming continuously in the tube as soon as this is in movement may be used. Because of this, fish living at the bottom of the sea and normally resting upon it must be omitted such as, e.g. the Greenland bullhead. Of the species studied, cod. five-bearded rockling and stickleback appeared equally suitable. Probably most Gadus species behave similarly, as well as species of other genera with a similar way of life. Cod as an experimental animal has many advantages, e.g.. its large range, the facility of obtaining regular catch from comparatively small depths, and its great resistance to the strain revolved in the present experimental technique. As is obvious from the reported observations on other fish. several species may be used as experimental animals.

Rotatory-riow technique applied to cod a

b

c

d

•

f .

g

.

.

.

h

~,35

In experiments with repeated test series the fish were brought back to the aquaria immediately after the last test in each series. The statements in the Tables on the temperature at the catch stations refer to the surface water. On the whole the temperature in deeper layers of the sea in the area varies in such a way that from March to October the temperature decreases with depth whereas the reverse prevails from October to March. A more detailed description is unnecessary' since we do not know the depth from which the fish invade the shallow water. With few exceptions all experiments were carried out at temperatures differing less than 3.0 C from that of the surface sea water at the catch station. In experiments with roach with the original version of the apparatus acceleration was 20. later 30 rev rain -2 (Lindahl and Schwanbom, 1971a. b~. As standard acceleration in most of the present experiments we have used 70 re~ m i n - ' . When not otherwise stated "'fish-length" means "'standard length", the total length minus the length of the caudal fin.

Behariour ql the cod in the rotational tubes during acceleration Fig. 2. Behaviour of cod in the rotating tube during different phases of the test. To the left the fish is seen from the side. to the right from above. (a and bt before rotation is started: [c and dl the fish still keeps its position with the head near the left wall of the tube but is inclined about 45 ° to the right; (e and I3 the head and the tail passively describe circular courses, the sagittal plane of the fish more or less coincides with the horizontal plane: (g and h) the fish with the belly upwards just before it is forced to rotate. Continuous circle marks the rotation of the tube. broken circle circular movements of the fish. cf. the text. It should, however, be stressed that in each case a similar study of special experimental conditions must be accomplished, as has been done in the present investigation. When stored in aquaria each fish was kept in about 401. of seawater, which was renewed at a rate of about 1 1. rain- 1. The water, passing a large tank, was taken from a depth of about 40 m. The salinity was about 33°,.... The experiments were performed at the Marine Biological Station of Kristineberg (Swedenl in the vicinity of which cod of suitable total length, 13-23 cm, are in abundant supply. The cod were caught with bow-nets at a depth of 1-3 m during the winter and 3-5 m during the summer, where the salinity varies between 20 and 28°00. The fish, generally living at greater depth during the day, invade these shallow waters at night. The bow-nets were set near the laboratory to avoid long transports and were emptied after one day and one night in order to eliminate the risk of poor condition due to an excessive stay in the bow-nets. The cod were then kept in aquaria with running water over night. Before the test. each specimen was transferred to the running water in a rotational tube of optimal width, to let the fish accustom itself to this environment until the increased frequency of ventilation caused by the handling etc., had decreased to normal values, and the fish did not show any signs of excitement. This generally required 30 rain. The fish was then tested 10 times in sequence with a 2 rain break between each test, The means of the results of such series were considered to be representative determinations of the critical rev rain-~-values (see p. 8411. * The observation of the fish from above was facilitated by a mirror, mounted above and parallel to the rotational tube on a level with the eyes of the observer and inclined 45 ° to the horizontal plane.

Before rotation is started the fish takes a position with the left side of the mouth region of the head against the right wall of the tube and the caudal fin near the left wall [Figs. 21a and bl]. In this position the head is lower than the tail. The movements of the caudal fin are exceedingly small or non-existent, while the pectoral fins slowly oscillate forwards-backwards, the right one now and then touching the wall. This position may be looked upon as the result of an avoidance reaction, as the fish probably perceives the observer, who is situated on the left-hand side of the tube. When the current of water starts rotating, the fish immediately reacts with increased activity of the pectoral fins. and soon also the caudal fin becomes more intensely involved by movements of the caudal muscles. The body is still oriented with the sagittal plane in a vertical position, nox~ with the head pointing obliquely towards the left and with the mouth region now and then touching the wall. Owing to this the left pectoral fin has space to move freely. With increasing rev rain- ~ the upright position of the fish is lost and the sagittal plane, inclined to the right, forms an angle of about 45: with the vertical plane [Figs. 2(c and d)]*. Characteristic for this phase are repeated efforts by the fish to bring the body into an upright position again, resulting in bending movements. Some time before the critical rev min-~ is attained the fish is forced into a position with the sagittal plane horizontal [Figs. 2(e and t)]. Figures 2(g and hi, show the fish with the belly upwards just before it is forced to rotate with the water [critical rev min-~). This is an irreversible and exactly defined event, independent of the time since the fish was forced to take side-position. The very moment when the fish just follows the rotating water mass round its own axis was chosen for switching off further acceleration and reading the critical rev min-~ on the tachometer. The ability to maintain the situation, shown in Figs. 2(e and ft increases with increasing current velocity: for some individuals it was observed that increase of the current velocit) from about 0.75-3.0 cm s- ~ resulted in an increase in the critical rex, min- ~ of about 15%. At the lower velocity the sudden rotation of the fish followed almost immediately after the commencement of the phase shown in Figs. 2(e and I) whereas the fish at the higher velocity could extend this phase for a longer time. thus attaining a higher critical rex' rain-1. In contrast this phase was reached at about the same rev m i n - : both at the higher and at the lower current velocity.

836

P.E. LINDAHL.S. OLOFSSONand E. SCHWANBOM

Dorsal and anal fins do not seem to be engaged in keeping the body upright. Whereas the pectoral fins are involved in steering and propelling movements, the chief power utilized by the fish for resisting the torque is obtained by muscles of the trunk via the caudal fin. With increasing rev rain- ~ the movements of the trunk become more violent and in the end phase appear as bendings of the whole body, i.e. when the tail fin is moved towards the left side of the fish. the head is moved in the same direction and vice versa. As the sagittal plane of the fish is more or less horizontal in the advanced phase of the test lFigs. 2(e and f)], the caudal fin beats up and down in the tube. At this the upward movement of the head Inear the left side of the wall) is supported by the rotating water, whereas the head in its downward movement beats against the rotating movement of the water. At the same time the head, moved passively in the direction of the rotating water, describes a circle. In most fish this circle falls more or less within the left half of the rotational tube. in others it fills out the whole tube. In this case the head intermittently touches the inside of the tube all around. Of course there are all kinds of intermediate stages between these two extremes. When the head describes small circles, the caudal fin also moves in circles, passing downwards near the right wall of the tube, then upwards through the central part of the tube where the rotatory rate of the water is low, and so forth. When the circles of the head become wider the same is observed for the caudal fin. No influence of the individual size of the fish on the behaviour during the test could be observed. During the whole course of a test the pectoral fins are frequently in touch with the wall of the rotating tube.

EXPERIMENTS

AND

RESULTS

To attain standardized experimental conditions the effects of certain biological factors must be investigated, such as the ability of the fish to familiarize itself with the strange surroundings in the rotational tube, optimal rate of flow, optimal relationship between fin-breadth (distance between extremities of pectoral fins measured perpendicularly to the length axis of the fishj and diameter of the tube, possible connection between size of the individual fish and critical peripheral velocity, and to what degree exhaustion or training affects the critical rev m i n with repeated tests during shorter or longer periods of time, and the effects of feeding in such experiments. The influence of temperature upon the critical peripheral velocity may be an important factor.

Familiarization After the transfer of the fish into the rotational tube it exhibits a behaviour suggesting a high degree of excitation. During the transport from the aquarium to the tube, as well as after this. the fish is exposed to different stress factors. We imagine that the very handling of the fish, the experience of finding itself in a limited space, the sudden water current and, in some cases, a change in temperature may be possible causes of stress. Excitation appears in attacks of the fish against the wall of the rotational tube or the inset-tubes at its inflow end, and in trying to turn downstream. These more violent reactions normally

end within a few minutes. In all cases the frequency of ventilation is increased by up to 100°~o. After about 30 min this frequency has become normal again and the excited behaviour has ceased. In exceptional cases this takes more time. After each repeated test in a test series the fish shows signs of excitement, but the return to normal behaviour generally takes less than 2 min.

Connection between individual size and critical peripheral velocity: "reaction quotient" With the introduction of different diameters of the rotational tubes, the critical rev m i n - t had to be replaced by some quantity, dependent of the radii of the tubes, viz. the critical peripheral velocity. This means that the critical rev m i n - ~ is multiplied by the interior circumference of the tube used for the determination. When critical pheripheral velocities were plotted against length of fish, linear correlation was revealed in a great number of experiments (Figs. 3-5, Tables 1-3, 5 and 6). As a provisional means of characterizing samples of fish by a figure which is independent of individual size, we divide the critical peripheral velocity by the length of each individual. The quotient obtained expresses the ability of the fish to react against the torque of the rotating water and is called "'reaction quotient". Assuming a fairly high degree of association between critical peripheral velocity and individual length in a sample of fish, and a regression line passing near the origin, the variance of its mean reaction quotient should be reasonably low. Calculation of reaction quotients implies greater variation when the regression line does not pass

~C

"~: 19001--

~

× ×~

•r.- 1 4 0 0

g

1300 o~,~.~ (J

14

;×

:

16

18 Length of fish,

:

,

20

22

cm

Fig. 3. Effect of varied water velocity upon the critical peripheral velocity, studied on a different sample of fish at each current velocity. For 15 of 30 specimens, caught at about the same time. the critical peripheral velocity was determined at 1.5 ( × j and for remaining 15 specimens at 3.0 (©~ cm s - 1 test series being performed alternatively at the two current velocities. For further information and statistics see Table I.

Rotator)-flow technique applied to cod "T c

~

22OO

o

o

• 2000

..~ 1800

"6 ,6o0 •~" 1400 1200 14

16 L.en(~4-h

20

18

of fish,

22

em

Fig. 4. Effect of varied current velocity upon the critical peripheral velocity, studied on a different sample of fish at each current velocity. For 10 of 20 specimens, caught at about the same time, the critical peripheral velocity was determined at 3.0 ( x ) and for the remaining 10 specimen at 4.5 (O) cm s- ~, test series at the two current velocities being performed alternatively. For further information and statistics see Table 2. through the origin or intersect the vertical axis near the origin. This will be discussed below (p. 843).

837

mean reaction quotient has increased by about 10%. At the same time the variation in the reaction quotients decreased, as demonstrated by the coefficient of variation (C/. Finally the mean range in critical rev m i n - ; in the test series is reduced by about 33°0 by the change in current velocity. However, the variation in range (S.D.) does not decrease, which makes the relative variation (C) increase considerably here. Next the velocity step between 3.0 and 4.5 cm secwas studied in a similar experiment, only with the difference that each group consisted of 10 specimens. In Fig. 4 the critical peripheral velocity has been plotted against length of fish. With both current velocities the points are rather close to the regression line. This is reflected by the high and highly significant correlation coefficients (Table 21. The regression lines are very nearly parallel lregression coefficients nearly equal): their intercepts, both negative, are numerically large but differ only insignificantly. Also in this experiment the increase in current velocity has induced an increase in the mean reaction quotient (about 7.5°o1. In contrast this was not accompanied by a decrease in C as in the previous experiment, nor did the range in rev min-1 values in the test series decrease.

Effects of varied streaming velocity During 1972 determinations of critical peripheral velocity were chiefly performed at the current velocity 1.5 c m s - 1. Since the choice of this velocity was rather haphazard, it seemed justified to investigate more closely the effect of variation of this experimental condition. In this study we concentrated partly on the variation within the test series, partly on the size and the variation of reaction quotients. As a provisional measure of the total variation within test series we used the range of r e v m i n - I expressed in 0o of the maximum value (100. [Xma, -- Xm,,]/Xm~0. In one experiment the critical peripheral velocity was determined at current velocities of 1.5 and 3.0 cm s-~ for 15 different specimens in each case, all brought into the laboratory within a few days. Fig. 3 shows the regression of critical peripheral velocity on length of fish. The good adaptation of the points to the two regression lines is reflected by the high correlation coefficients (Table 1). The two regression coefficients b differ so much that the regression lines do not appear parallel. The difference in b is not statistically significant. Both regression lines intersect the vertical axis on the positive side of the origin. The

IC

2ooo _; ,,°° ,4°°rI-

3,

,2oo r 0

14

16 18 Lenqth of fish,

20 cm

22

Fig. 5. Effect on the critical peripheral velocity of varied diameter of rotating tube in relation to fin-breadth. For each one of 8 specimens (length 14.6--22.0cm) critical peripheral velocities were determined in three different situations: (1)the diameter of the tube exceeded the fin-breadth by more than 5 mm; (2) both measures were about equal (in three exceptional cases the tube diameter was 5 mm shorter than the fin-breadth), and (3) the fin-breadth exceeded the tube diameter by 10 ram. The current velocity was 3.0 cm s-L For further information and statistics see Table 3.

Table 1. Comparison of results obtained at two current velocities with two different groups of 15 individuals in each (cf. Fig. 3). Rotating tube diameter adjusted to fin-breadth. Length of fish at 1.5cms -t 13.1-21.8cm, at 3.0cm s -t 13.2-21.5. February 1975: Temperature at catch station 4C, of aquaria 6-7~C and at testing 5~'C Current veJocitx cm s- 1 1.5 3.0

Correlation between critical peripheral velocity and length of fish r t P h 0.96 0.98

12.36 17.76

<0.001 <0.001

* Difference 9.2, t = 6.91. P < 0,001. 4-Difference 4.3, t = 7.07, P < 0.0(31.

~().2 92.1

Mean reaction quotient

IRQI

Inlercept, cm rain i

R--Q _-'-.S.D.

CI%)

8q 32

85.3* ~ 4.23 94.5* _. 3.95

50 3,1

Mean range (R) of rex min-~ ,,alues in test ~eries. (%) R _+ S.D. C ('!,) 13.07 +_ 3.68 8.7~" + 3.62

28.3 44.9

838

P.E. LINDAHL, S. OLOFSSON and E. SCHWANBOM

Table 2. Comparison of results obtained at two current velocities with two different groups of 10 individuals in each (cf. Fig. 4). Rotating tube diameter adjusted to fin-breadth. Length of fish at 3.0cms -~ 13.6-21.5cm. at 4.5cms -~ 14.5-21.5 cm. May 1974: Temperature at catch station 13.1 :C of aquaria 9.5:C and at testing 10.OC Current ',elocity c m s -I 3.0 4.5

Correlation between critical peripheral ~elocity and length of fish r t P h 0.97 0.92

11.28 6.64

<0.001 <0.001

Intercept. cmmin -t

107.2 109.8

--376 --311

Mean reaction quotient (RQI ,~Q X S.D. C (l'r~ 86. l* + 5.17 92.6* ± 655

6.0 7.1

Mean range IRI of ~a[ues m test series t",,) RxSD C I"ol 143+ Z 4.58 12.8+ + 2.94

32.0 23.0

* Difference 6.5. t = 2.46, P < 0.05. t Difference 1.5, t = 0.89. P < 0.4 Table 3. Comparison of results obtained in different situations involving varied space surrounding the fish in the rotating tubes (cf. Fig. 5). Test series performed with identical fish (length 16.3-22.0cm) on three successive days in the order shown below. January-February 1974: Temperature at catch station Y-C, of aquaria 4 C and at testing 5.0~C. Tube diameter exceeding fin-breadth

N u m b e r of fish

Correlation between critical peripheral selocity and length of fish r t P b

Intercept. cm r a i n - t

Mean reaction quotient IRQI Significance of differences RQ ± S.D. C I",0 t

>5

7

0.71

2.25

<0.1

83.9

140

91.46 + 6.02

1036

-0

8

0.91

5.38

<0.01

86.2*

- 125

7964 _+ 550

738

-10

8

0.86

4.13

<0.01

64.1'

59

67.54 ~- 5.12

8.08

2.97

<0.01

426

< 0.001

* Difference 22.1. t = 2,111, P = 0.1. Table 4. Comparison between first-test series performed during the periods 1972 and 1973-74. i.e. before and after the introduction of the net behind the fish, the increase in the standard current velocity from 1.5 to 3.0 cm s-~. and the adjustment of the rotating tube to the fin-breadth. For definition of range of critical rev min- ~ see text p. 837. Inclination refers to that of the regression of critical rev rain- ~ on serial number of tests: Length of fish varied from 13 to 23 cm, temperature of tests from 5 to 15~C

Period of time 1972 197374 Significance of differences

No. of test series 28 35 f t ~ P

Mean range I%1

Distribution of test series I",,I Inclination of regression not significant

19.0 103 6.()0~ < 0.001

85.7 71.4 1.41~ < 0.2

Inclination of regression significant Regresston Regression positive negative 36* 213.(r~ 2.t 6~ < 0.05

[0.7" ~6+ I).27~ < 0.8

* Difference 7.1, t = 1.04, P < 0.4. "I"Difference 11.4 t = 1.38, P < 0.3. Students t-test t for comparison between found proportions (Bonnier and Tedin. 1957, p. 51: cf. Garret. 1945). Summing up, we find that raising the current velocity causes increase in the mean reaction quotient. In the velocity step 1.5-3.0cms -1 this is accompanied by a decrease in the variation of reaction quotients a n d rev m i n - 1 values in test series, but this is not the case in the step 3.0-4.5 cm s - ~. The advantage of higher values of reaction quotients in the latter case does not compensate for the drawback of less exact readings of critical rev min-~-values on a less favourable scale of the tachometer. Accordingly 3.0 cm s-~ was chosen as standard current velocity.

Effects of varied diameter of rotational tube It is easily realized that a small fish in a wide rotating tube, residing in its central part, will not be influenced by the high rotational velocity of the peripheral water masses in the tube. On the other hand a large fish in a narrow rotational tube will not have enough space for efficient use of its caudal and pectoral fins. One may except too high a critical rev-

m i n - 1 in the former situation and possibly too low a critical rev m i n - 1 in the latter. In order to test the validity of this conception the critical rev m i n - ~ for 8 fishes were determined on 3 successive days, starting on the first day using rotating tubes with diameters exceeding the pectoral finbreadth (cf. p. 836) of the fishes by at least 5 mm. O n the second day the tube diameter and the finbreadth were very nearly equal, except in three cases where the diameter was 5 m m shorter than the finbreadth. O n the third day the tube diameter was in all cases 10 m m shorter than the fin-breadth. In all cases the current velocity was 3.0 cm s - t . Generally the critical peripheral velocity of each fish increased with the space at its disposal. This appears also from the arrangement of the regression lines in Fig. 5 in which the single experimental points have been omitted. The results of the statistical treatment have been collected in Table 3. W h e n the diameter of the rotational tube exceeded the fin-breadth by more than

Rotatory-flow technique applied to cod

839

Table 5. Effects of repeated test series performed at the current velocity 3.0 cm s-~. The fish of experiment I were fed. those of experiment 1I and III not. Temperature was in experiment I 14:C (at the catch site 12~CL in experiment II 1 5 C [at the catch site 15:Ck and in experiment III 6.OC lat the catch site 5.5°C): the number of fish was in experiment 1 15. in experiment II 15. and in experiment III 10. Standard length of fish was used in version la) of experiment I and in experiment III. total length in version (b) of experiment 1 and in experiment 11. Diameter of rotating tube adapted to fin-breadth in experiments I and II. In experiment III the ratio between length of fish and tube diameter was about 2.36. Experiment 1 and II:. October. experiment II December 1974.

Experiment

Length of fish. cm

lla) 13 4-21.0

l(bl 14.6 228

II 14.3-23.0

111

13.8-21.4

* Mean + Mean Mean § Mean

difference difference difference difference

Correlation between critical peripheral velocit 3 InterN o of test and length of fish cept series r b tom m i n - t l

Mean reaction quotient (RQI RQ +_ S . D C Col

1

0.90

92.1

-15

91.2" 4- 2.23

2.4

2

0.06

066

-98

9 0 6 ~ 370

41

3 I

094

0.99

87,8 86,2

31 -37

89.7" 4-_ 3.08 84.1";" 4- 2 0 0

5.0 2.4

2

096

90.5

-122

83.6 +_ 3.38

4.0

3 I

0.95 0.06

82.6 96.6

4 -188

82.85" +_ 3.94 86.4~ + 5.24

4.8 6.0

2

0.93

92,2

-140

84.6 + 5.80

6.9

3 1

0.95 0.97

92.2 98.7

-180 50

82.5~ ~ 5.12 101.6~ Z 4.49

6.2 4.4

2

096

046

133

102.3~ _+ 5.67

5.5

3

0.95

89.8

170

99.7~ ~ 5.32

5,3

-1.53; -1.35; -3,93; -2.02:

f -t= t= t=

1.86: P 1.78; P 4.62; P 3.61 : P

< < < <

Ditlerence between reaction quotients Deviation of mean from zero Mean t P

-0.63

0.80

<0.5

--09I

1.06

<0.3

-0.49

0.64

<0.6

--0.79

1.01

<0.4

-- 1.83

1,78

<0.1

-2.10

2.90

<0.02

- 1.02

1.57

<0.2

- 2.76

3.07

< 0.02

0.I. 0.1. 0.001. 0.01.

5 mm there was no significant correlation between critical peripheral velocity and fish length (P _< 0.1). In contrast the correlation coefficient was significant both when the tube diameter and fin-breadth were about equal and when the tube diameter was 10 mm shorter than the fin-breadth. The regression coefficient in the latter case is smaller than that in the former. The difference is, however, not significant. On the other hand the mean reaction quotient was significantly greater when fin-breadth and tube diameter

were about equal than when the fin-breadth exceeded the tube diameter. In the latter situation the fish may not have sufficient space to use its fins effectively. Disregarding this poorly substantiated explanation, it seems appropriate to aim at high measurement figures as long as this does not involve any disadvantage. Thus the situation with about equal tube diameter and fin-breadth is the most ideal of those studied in this experiment, and we arrive at the conclusion that the tube diameter must not exceed the fin-breadth

Table 6. Effects of small changes in temperature on reaction quotients. The three test series of each experiment were performed ~ith the same specimens on 3 successive days in the order given below. The current velocity was 3.0 cm s -t and the rotating tubes were adjusted to the fin-breadth. Experiments (A and B1 August 1974; temperature at catch station 16.5:C. Experiment C: March 1975, temperature at catch station 3°C, of aquaria 6°C. The fish of experiment C were offered food daily

Experimem

A .

B

C

Number of fish (length of fish. call

15 112.0-21.5t

14 il2.tL 18.0i

20 I11.8-21.1 )

Temperalure of testing

Correlation between critical peripheral velocity and length of fish Intercept, r h cm rain-~

Mean reaction quotient (R--Q) R'Q ± S.D. C [°~,l

13

0.97

Q6.5

-96

90.5* _-'24.64

5.13

16

0.98

95.3

-97

91.2 -L-_4,49

4.92

19 16

0.98 0.88

98.3 97. "~

-151 -60

89.6" + 4.44 938¢ ~ 7.25

4.96 7.73

19

0.8"7

90.1

93.0 ~ 6.67

7.17

44

22 5

0.88 0.97

03.0 04.5

-I0 - 31

92.3* _+ 7.06 92.5,* -4" 4.74

7,65 5.1

I1

094

94.0

-35

91.7 4- 6.17

6.7

8

0.96

991

* Mean difference -1.08, t = 1.00, P < 0.4. -1-Mean difference -1.49, t -- 2.86, P < 0.02. Mean difference 0.41, t = 0.66, P < 0.6.

-112

92.1~ ± 6.28

6.8

Mean difference

Deviation from zero t

P

0.71

1.16

<0.3

- 1.83

1.49

<0.2

-076

1.15

<0.3

-0.66

1.20

<0.3

1.09

1.68

<0.2

0.26

0.32

<0.8

840

P.E.

LINDAHL S. OLOFSSON a n d E. SCHWANBOM

by more than 4 mm. The experiment presented in Table 3 was also analyzed with respect to the range of rev m i n - 1 within test series. The lowest mean range appeared when diameter of rotating tube and finbreadth were about equal. Only one fish out of 8 (7) had in this situation a range larger than in the two other situations. However, the variation in range in each situation was very large (C = 35--49°0), and the means did not differ significantly. " Comparison bet~,een ran#e within test series of early and recent determinations of rev min- 1 In the two previous sections the influence of changes in current velocity and in the space surrounding the fish upon the variation of critical rev min-1. values was studied. For practical reasons the number of fish was rather small in these studies. It therefore seemed necessary to evaluate in some way the combined effects of these methodological changes, including also those due to insertion of nets behind the fish. This was done by comparing the range of rev m i n - ' - v a l u e s in test series in some different respects. Thus series performed before and after the introduction of the mentioned changes were collected at random, i.e. from the two periods 1972 and 1973-74. Each of all these determinations of mean critical rev m i n - L v a l u e s was the first one performed with the specimen in question (first-test series). As is shown by Table 4 there is a striking decrease in mean range

T_E r

"Xj

: "u-'\

<'>' /',,

: 2

' 3

1 4

I 5

I 6

i 7

': -> , o d \

x.-.-x....-.-x (a I

8°1-

of about 46°.0 between the two periods. As is obvious from Fig. 6 three types of curves relating critical rev m i n - ~ to .serial number of test, may be distinguished, viz. (1) such with neither upward nor downward tendency, (2) such with upward, and (3) such with downward tendency. Within these three types, which, of course, are not sharply delimited, there is a great variation as to the dispersion of points. Thus some curves show a smooth course whereas others are more irregular. Some still more irregular than those demonstrated in Fig. 6 have occurred. Because of these irregularities we desist from defining certain slopes, partitioning groups 2 and 3 from group 1. Instead the correlation coefficient (rl was calculated for all series, and all cases in which r differed significantly from zero were taken to group 2 or group 3 according to whether the regression of critical rev m i n - 1 on serial number of test was positive or negative. Remaining cases were classified as belonging to group 1. In both samples of curves those lacking significant inclination dominate, constituting 85.7 and 71.4°,o of those collected from 1972 and 1973-74, respectively (Table 4). The difference is not significant. This means that the majority--about 78°o----of the fish are not affected during first-test series. The distribution of the remaining cases between groups 2 and 3 does not involve a significant difference in either of the two samples of curves. Thus we have no reason to believe that the proportion between

I 8

J 9

t3

,

7o ~ " x

I IO

t ~

N u m b e r of test

/\ I 2

l 3

', 4

5

6

, 7

' 8

, 9

iO

Numimer of test

Fig. 6. Types of curves of test series selected at random. Type 1, neither upward nor downward tendency, a, b, c. Type 2, upward tendency, d. e. Type 3. downward tendency, f, g. The examples are chosen so as to demonstrate different ranges within series of rev rain- i. The curves were obtained under the following conditions: Test series a b c d e f g

Length of fish 19 15 20 20 25 23 18

t tube dia (mm) 5 10.5 II 5 10 8 11

70 50 60 70 80 70 55

Current velocity, (cm s- 1) 3.0 1.5 1.5 3.0 3.0 1.5 1.5

Correlation coefficient r 0.57 0.03 -0.I1 0.97 0.68 -0.76 -0.68

Only when the current velocity was 3.0 cm s- 1 were nets used behind the fish.

Range of critical rev min- ~(°o) 3 10 20 13 10 23 18

Rotatory-flo~x technique applied to cod test series with positive and negative regression, respectively, was influenced by the changed experimental conditions. Summing up, it is obvious that these changes have brought about a marked decrease in the source of error in the determination of critical rev min- 1.

Reproducibility of reaction quotients and the effects of repeated tests Inspection of series of critical rev min-1 obtained in test series performed under somewhat varied experimental conditions reveal great variation, the figures of critical rev rain-~ when plotted in successive order forming curves of varying appearance (Fig. 61. Although there may be a great variation within each series, one is given the impression that the greater part of the curves show neither upward nor downward tendencies (cf. Table 4}, This suggests that the fish during series of 10 successive tests, neither "learn" how to resist the strain of the testing procedure better, nor do they grow increasingly more tired during the test series. However, since the values may be rather scattered it was thought necessary to calculate mean values from as many as 10 tests (cf. p. 835). It is of great interest to check the reproducibility of the method. At the same time it is of importance to know possible effects of frequently repeated test series. For this purpose one test series was performed at a current velocity of 1.5 cm s-1 each day during 5 successive days with 14 cod of varied body length (13-25cm). Mean reaction quotients showed a decrease, but none of the means of day 2 to 5 differed significantly from that of day 1. The coefficient of variation of the mean reaction quotient increased with the number of test series, showing that the fish were in some way influenced. However, this experiment was carried out before the standardization of experimental conditions had been accomplished, when the reactions of a studied sample were more heterogeneous. Therefore, the effects of repeated test series were studied at the current velocity 3.0 cm s(Table 5), including 3 test series on 3 successive days. The effect of feeding was investigated in experiment I and it was also examined whether the total length icf. p. 835) of the individual is a better expression of its size in calculations than the standard length (exp. I. version (bt experiment II). Therefore, when comparing experiment II with experiment I, version I (b) should be consulted. Experiment III may be directly compared with experiment I(a), although the relationship between tube diameter and fin-breadth was a little different (cf. Table 51. The main result of the experiments in Table 5 is that in all three cases no significant change in reaction quotient can be detected between the first and the second day, although variation is kept on a reasonable level. Furthermore, in two out of three cases ~experiment II and III) the mean reaction quotient of the third day is significantly lower than that of the * Student's t-test on estimated variance of difference between intercepts.

841

first day. The corresponding lack of decrease in experiment I may be due to the fact that the fish had been fed, and this idea is further strengthened by the result of experiment C in Table 6. In some instances certain statistics are regularly changed throughout an experiment, although without statistical significance between successive values. The continuously increasing intercept in experiment III of Table 5 is of great interest: however, it is not significant*. Referring critical peripheral velocities to the total length of the individual fish instead of to their standard length (Table 51 appeared chiefly as an 8°o decrease in reaction quotients and as a displacement in the negative direction of the points of intersection of the regression lines on the ordinate (Table 6t. No striking changes in variance could be observed. The rather high reaction quotients in experiment III are due to the different relationship between tube diameter and length of fish. An attempt to trace a connection between the variation in the reaction quotients of the individual fish and the variation in the corresponding test series of the experiments with repeated tests gives the following somewhat generalized picture. The number of fish showing a clear upward or downward tendency in the reaction quotients of the three days is comparatively small. When the reaction quotient of the second day in such cases is decidedly lower than that of the first day, this is generally caused by a considerable increase in the test curve of the first day making the corresponding mean reaction quotient especially high. The reverse state of affairs occurs when the mean reaction quotient of the second day is decidedly higher than that of the first day, the test series of the first day showing a considerable decrease. Strikingly great changes in the test values generally occur on the first day and are rare on the following days. Besides, the variations within the test series generally decrease and are inconsiderable in the second half of the series. However, the sums of the first 5 and of the later 5 tests do not differ significantly. Also the origin of the difference in display of the mean reaction quotient in experiments with unfed and fed fish should be traced back to the reaction quotients of individual fish. In the former case (Table 5, experiment II) 8 fish out of 15 show a significant decrease in the reaction quotient between the first and the third day, whereas in the latter case (Table 5, experiment II significant decrease occurred in 4 out of 15 fish. Decrease in mean reaction quotients obtained on successive days might thus be looked upon as a consequence of starvation (cf. also Table 6. experiment C).

Effect of changed temperature The determination of reaction quotients, standardized to be performed at certain fixed temperatures near that of the sea water at the catch station infers that the temperature of the environment of the fish

842

P.E. LINOAHL.S. OLOFSSONand E. SCH~'ANBOM

is usually changed by 2-3°C, We have therefore carried out control experiments in which the fish were exposed to temperatures 2-3°C below and above that at the catch station, and in some experiments also to a more deviating temperature. In all cases the same specimens were studied at all three temperatures used in the experiment in question, in order to minimize the effect of variation between individuals• This means that each individual was submitted to test series repeated during 3 successive days. Experiments were performed at current velocities of 1.5 and 3.0 cm s - 1 In the former case increases in temperature of 3-4~C (initial temperature 5'C) did not cause any change in mean reaction quotients, whereas increases of 6-1OC ithird-test series) induced decrease. Since the variance of the mean reaction quotients was high from the beginning and increased further with the number of test series performed, as in other experiments of similar kind (cf. p. 841). we desist from any conclusions. Three experiments were performed at the current velocity 3.0 cm s-1 (Table 6, IA-C)). In experiment A the temperature at the catch station was 16.5:C. The medium temperature was 16.0°C, i.e. about the same as that at the catch station, whereas the lowest and the highest temperatures differed from the medium by 3.0 °. At all temperatures the critical peripheral velocity correlated well with the length of the fish. The regression coefficients differed very little. Nor did the means of reaction quotients differ significantly in spite of the rather low variation (for coefficients of variation see Table 6). The same result appeared when the significance of mean differences between reaction quotients of individual fish were analyzed. This means that neither the decrease, nor the increase in temperature had any effect on the reaction quotients in this sample of cod. In experiment B of Table 6 the lowest temperature was about equal to that of the sea water at the catch station. It was increased in two steps of 3.0:C. Also in this case correlation between critical peripheral velocity and length of fish was highly significant, and the regression coefficients differed insignificantly. The mean reaction quotient decreased continuously but not significantly. An analysis of this decrease by applying a test of significance to the means of differences from individual fish indicates that the change in temperature from 16 to 1 9 C did not affect the mean reaction quotient, whereas the change from 16 to 22~C brought about a rather small but significant decrease. Since this decrease, appearing in the third of 3 repeated test series, is of about the same magnitude as that observed in experiments on effects of repeated test series (see abovel, it is doubtful whether it is an effect of increased temperature. * Values of reaction quotients obtained from third day test-series and probably too low, since the fish were not fed, cause too ~eat differences between experiments in Table 3 itube diameter and fin-breadth about equal and tube diameter 10 rnm shorter than fin-breadtht and Table 6B Itemperature 16 and 22 C).

In experiment C the lowest temperature (5°C), studied on the first day, was 2:C higher than that of the sea water at the catch station. On the second day the test temperature was 1 I:C and on the third day 8:C. In this way an increase in temperature of 6~C was inserted between the first and the second testseries, i.e. without any risk of invalidation of this comparison by effects of repeated tests. All of the 20 fish used in this experiment were caught on the same site on the same occasion. Because of the great number of fish, the experiment went on for 14 days. During the period of time from the catch to the last test the fish were offered feed daily. The correlation of critical peripheral velocity and length of fish was high in all the three test-series. The three regression coefficients were rather simi!ar, only the one for 8 C differed somewhat [difference not statistically significant). The intercept was subject to a similar change which was, however, not statistically significant. The mean reaction quotients did not differ signifi/:antly when compared directly, nor when the mean differences were tested for deviation from zero. DISCUSSION In order to make possible experiments also with species in which the quotient between largest frobreadth or height and the length is lower than that in the cod, the rotational tubes were made longer than necessary for the present investigation. Even when the fish is kept as near the inflow end of the tube as possible with the aid of a net behind it (p. 834) an effect of the size of the fish on the variation of the results is inevitable. A correction based upon this fact is so small that it may be neglected. We have not been able to detect any influence of changed current velocity on behaviour, position, or orientation in the tube of the individual specimen during the test. We have therefore advanced the hypothesis that it is easier for the fish to resist the torque when the rate of its forward-swimming is greater. This might possibly also be the explanation for the greater stability of its reaction throughout a test series at 3.0 than at 1.5 cm s - t . It provides a great advantage if a sample of fish can first be used as control sample and then, after some treatment, function as experimental sample. In such a case the effect of the variation between individuals may be reduced, when testing the significance of the difference, by calculating the mean difference from the differences of individual fish, and test its deviation from zero. A condition for this procedure is, however, that repetition of test series on successive days does not influence the reaction of the fish. This was controlled in experiments performed at the current velocity 3.0 cm s- 1. The only effect observed was a small but significant decrease in the reaction quotient on the third day in two out of three experiments. The nutritional state was suggested to be a contributory cause of this decrease*,

Rotatory-flow technique applied to cod Within the range of temperatures tested, 5-22 C. changes of 3:C did not cause any change in reaction quotients, neither did changes of 6:C unless too high a temperature was attained (Table 5, experiment C). The upper lethal temperature of cod is stated to be +20~'C (Wise, 1948). The general conclusion of these experiments is that testing may be performed at temperatures in the vicinity of that of the habitat of the fish, and that there is no need to use this exact temperature, The cause of the range of critical rev min- ~ in test series is two-fold. The one component depends on stocastic variation and is the only source of variation in series devoid of inclination, whereas the other component is a consequence of inclination• A decreased range thus implies less variation in the determination of critical rev rain -1 and mean reaction quotients. The improved experimental conditions resulted in a fairly large decrease (Table 4) of about 45°o of the range in test series• The major part of this change is caused by the increase in the current velocity from 1.5 to 3.0cms -1 (cf. Table 1}. Also the adjustment of the tube diameter to the fin-breadth was accompanied by a decrease in mean range (cf. p. 840) which was, however, not statistically significant. Rheotaxis is a prerequisite for rotatory-flow technique. We have not made any detailed study of the role of visual stimuli in induction of rheotaxis (Lyon, 1905) in our experiments, However, it was shown by Harden Jones (19631 that cod. which in free water orients to a background coming up at its sides from behind with velocities corresponding to 1-2 cm s-1. fails to do so when at the bottom. In keeping with this we have not been able to induce any marked forward-swimming of the fish in the tube, before rotation had started, by applying a moving striped background at one side of the tube. Tactile stimuli were shown by Dykgraaf 11933) in minnows to be of great importance for rheotaxis. The frequent touching of the wall of the rotating tube with head and pectoral fins by the cod (p. 835) provides the fish in the apparatus with tactile stimuli in abundance. From the point of view of the fish it must be immaterial whether it is sliding past the firm surface because of the streaming of the water, or whether the former is moving in relation to the fish (rotation of the wall surfacel. The fish is determined to swim in the direction opposite to the resultant of these two velocities (Fig. 7). This velocity resultant soon forms a relatively large angle with the axis of the rotational tube. Thus the fish, touching the right side of the tube, may soon become inclined to move upwards and then, touching the upper part of the tube, to pass over to the left side. Here the wall of the tube comes towards the fish chiefly from below, inducing the fish to turn its head downwards. In this position the left region of the snout and the left pectoral fin touch the wall slightly below the plane through the horizontal diameter, where the direction of motion is chiefly

843 j"7 f'7

J

Vr4~

Vf2

Vs2

Velocity of

rotation

Fig. 7. Velocity diagram projected on the interior surface of the rotational tube. showing the influence of two different current velocities (Vf~, Vf2) upon the velocity (direction) of swimming. Low current velocity IVfl) gives the angle :~, high current velocity (Vf,) the smaller angle ~'2. Vr~ and Vr2. velocity resultants: Vsl and I/s2 corresponding velocities of swimming.

upwards. Since the ventral parts of the fish do not touch the tube, no sensations of the motion from right to left below the fish are perceived, and the fish will strive to remain in this situation and at the same time maintain its upright position. When in a later phase of the test it inclines more and more towards the right and describes circular motions, this seems to be caused by the rotating water. When we proceed from these qualitative considerations to an explanation of the quantitative changes, resulting from changed current velocity, greater difficulties are encountered. The direction of swimming induced in the fish is at any moment in the opposite direction to the resultant of the current velocity and the velocity of rotation (Fig. 7). Consequently, the swimming is from the beginning directed against the inflow end of the tube, and, as rotation starts and the rotational velocity increases, the angle (a) which it forms with the axis of the tube gradually increases. Since we are striving towards a reduction of the variation within samples of reaction quotients, it is of interest to inquire into the genesis of the intercepts with the present manner of performing the experiments. Since both pectoral fin-breadth (-) and length of fish (x), here expressed in cm. are involved, we start with the relationship between both and find, by measuring 122 cod with the standard length ranging between 13.4 and 24.3 cm, a rectilinear correlation - = 0.41x + 0.06 (coefficient of correlation 0.98). The interior diameter of the rotational tube exceeds the fin-breadth by

844

P . E . LINDAHL. S. OLOFSSON and E. SCHWANBOM

about 0.4cm. The interior circumference (cl of the rotational tube is then c = rcl0.41x + 0.06 + 0.4) The interior circumference multiplied by the critical rev m i n - i (vc) gives the critical peripheral velocity (yk y = t'c.i'r~.(O.41x + 0.06 + 0.4)

(i individual coefficient of fitnessl. If ),, being "too large" in relation to x with a constant number, is plotted against x the straight line obtained will intersect the vertical axis on the positive side of origo. In spite of this the majority of the regression lines resulting from the present experiments intersect the vertical axis on the negative side of origo (16 out of 21), the first series in Table 3, experiments I(b) in Table 4, and III in Table 5 being excluded for different reasons from this grouping). This suggests that some factor other than those considered in the formula is operating. A survey of the mean reaction quotients of the 7 control samples of cod presented in the Tables of the present investigation shows a variation from 86.1 to 94.5 (9.7°/0) and a mean of 89.91. There is no connection between mean reaction quotient and temperature. It thus seems probable that different samples of fish differ in fitness because of variations in one or more unknown factors. At a properly chosen current velocity the critical peripheral velocity is linearly correlated to the length of fish. From a physiological point of view this relationship does not seem probable, since from purely theoretical considerations the maximal force a muscle can develop is proportional to its surface and thus proportional to the second power of the length of the individual, whereas the work performed should be proportional to the third power of the length (cf. Hill. 1950; Astrand and Rodahl, 1970). It would be of great interest to be able to calculate the force acting on the fish in the direction of rotation just when the critical rev rain-~ is attained. However. hydrodynamic theoretical studies concerned with immersed bodies moving through a medium, or stationary in a moving medium, are generally directed towards calculation of the resistance which the body encounters or the force that the medium exercises on the body in the direction opposite to that of the motion (cf. Rouse. 1961: Daily and Harleman, 19661. As far as we are aware the theoretical prerequisites for calculating the force of interest in our case have still not been developed. We are therefore constructing a device allowing measurements of the force exercised by the revolving water on models of fish. SUMMARY

1. When the current velocity was increased from 1.5 to 3.0 and further to 4.5 cm s- ~ the mean reaction quotient increased significantly in both steps. At the same time the mean range of critical rev min-~ in

the test-series of the different specimens decreased in the former step but not in the latter, indicating a reduction of variation in the former step. A standard current velocity of 3.0 cm s- t has been used. 2. With this current velocity the effect of varied relation between rotational tube diameter and finbreadth was studied, using the same fish on three successive days. The wider the tube the higher was the mean reaction quotient. With the tube diameter exceeding the fin-breadth by more than 5 mm there was no significant correlation between critical peripheral velocity and length of fish: this situation was accordingly rejected. With tube diameter and finbreadth equal and with the fin-breadth exceeding the tube diameter by 10 mm, correlation between critical peripheral velocity and fish length was significant. The mean reaction quotient was significantly higher in the former than in the latter situation. The tube diameter was made exceed the fin-breadth by not more than 4 mm as a provisional standard. 3. First-day test-series were selected at random from experiments performed (A) before and IB) after increase of the current velocity from 1.5 to 3.0 cm s- 1, introduction of detailed adaptation of tube diameter to fin-breadth and of nets behind the fish. Comparison of results from ~A) and (B) shows a striking decrease of about 46°0 in the mean range of rev rain- 1 in test-series. In both cases about 78"0 of the testseries lack significant inclination, indicating that the majority of fish were not affected during first-day testseries. 4. In experiments consisting of 3 test-series repeated on 3 successive days at a current velocity of 3 . 0 c m s - l , no significant change in mean reaction quotient between day 1 and day 2 could be detected. In two experiments out of three there was a significant decrease between day 1 and 3. In the deviating experiment the fish had been fed. in the other two not (cf. also Table 6. experiment C). Feeding is therefore recommended in co.nnection with this kind of work. 5. Changes in temperature of 3 - 6 C induced significant changes in mean reaction quotients only when the upper critical temperature of cod was reached. 6. With the rotating tube diameter exceeding the fin-breadth by not more than 4 mm, the regression of critical peripheral ~elocity on fish length should, from theoretical considerations, be expected to intersect the vertical axis on the positive side of origo. In spite of this the point of intersection is found on the negative side of origo in the majority of test-series ~16 out of 21 in the present reportl. Acknowledgements--This study was financially supported by the National Swedish Environment Protection Board. The new version of the rotatory-flow apparatus was skilfully designed and produced by Mr. E. Nyberg. Our thanks are due to Dr. B. Swedmark and the laboratory staff of Kristineberg Marine-biological Station for facilitating our work. We thank Mr. G. Steinholtz for valuable technical discussions.

Rotatory-flow technique applied to cod REFERENCES

Bengtsson B. E. (1975) The effect of zinc on the ability of the minnow, Phoxinus phoxinus L.. to compensate for torque in a rotating water current. Bull. Environ. Contain. Toxicol. 12, 654-658. Daily J. W. & Harleman D. R. F. (lq661 Fluid Dynamics. Addison-Wesley. Reading. Mass. Dykgraff S. (1933) Untersuchungen tiber die Funktion der Seitenorgane an Fischen. Z. vergl. Physwl. 20, 162-214. Harden Jones F. R. (19631 The reaction of fish to moving backgrounds. J. expl. Biol. 40, 437~,46. Hill A. V. 11950) The dimensions of animals and their muscular dynamics. Proc. R. lnsm. G. Br. 34, 450-473.

845

Lindahl P. E. & Schwanbom E. (1971at A method for the detection and quantitative estimation of sublethal poisoning in living fish. Oikos 22, 210-214. Lindahl P. E. & Schwanbom E. (1971b) Rotatory-flow technique as a means of detecting sublethal poisoning in fish populations. Oikos 22, 354-357. Lyon E. P. (1904) On rheotropism. I Rheotropism in fishes. Am. J. Physiol. 12, 149-161. Rouse H. (1961) Fluid Mechanics for Hydraulic Engineers. McGraw-Hill. New York. Wiese J. P. 119581 Cod and Hydrography--a review. Spec. scient Rep. U.S. Fish Wildl. Serv. Fisheries. (245} 16 pp. Astrand P.-O. & Rodahl K. 11970~ Textbook of Work Physiology. McGraw-Hill. New York.