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Oxygen consumption of the Cladoceran, Daphnia pulex, as a function of body size, light and light acclimation
Camp. Biochem. Physiol., 1972, Vol. 42A, pp. 877 to 888. Pergamon Press. Printed in Great Britain
OXYGEN CONSUMPTION OF THE CLADOCERAN, DAPHNIA PULEX, AS A FUNCTION OF BODY SIZE, LIGHT AND LIGHT ACCLIMATION* ARTHUR
L.
BUIKEMA,
JR.~
Department of Zoology, University of Kansas, Lawrence, Kansas 66044
(Received 15 November 1971)
The effects of nine light intensities, polarized light and four wavelength ranges were studied on the metabolic rate of Duphnia. 2. The effect of light intensity was significant and the effects varied depending on, light intensity, body size and state of acclimation. 3. Except for differences among adjusted means the effect of polarized light was inseparable from the effect of light intensity. 4. Acclimation to wavelength reduced metabolic rate and there were no other significant effects.
Abstract-l.
INTRODUCTION
LIGHT intensity affects vertical migration and changes in locomotory activity of Cladocera (Yerkes, 1900; Dice, 1914; Schulz, 1928; Clarke, 1930, 1932; Cushing, 1951; Berner, 1962; Ringelberg, 1963, 1964) and the response to light intensity varies with age (Clarke, 1932). Polarized light also affects the behavior of aquatic organisms and the effect has been described as one of increasing light intensity in the cone lenses (Baylor & Smith, 1953, 1957; Bainbridge & Waterman, 1958; Hazen & Baylor, 1962). Wavelength also affects activity and red wavelengths (620-640 rnp) are most effective in stimulating the activity of Daphnia pulex, D. magna, Leptodora kindtii and Moina. Lesser stimulatory wavelengths occurred around 540 rnp for Moina only and 440 rnp for the other species (Lumer, 1932). Blue wavelengths (400450 rnp) may be more stimulatory than red wavelengths (680-700 rnp) for Simocephalus vetulus and the shrimp, Paratya compressa, but there are no data for the 550-680 rnp range (Nagano, 1955). Smith & Baylor (1953) and Baylor & Smith (1957) report that blue and red dances are exhibited by cladocerans, copepods, fairy shrimp and SquiZZu larvae and that blue wavelengths distinctly agitate the populations so that the velocity of the blue dance is three to five times that exhibited in the red dance. The ostracod, Cyprinotus incongruens, was more * This paper is part of a Ph.D. thesis submitted to the Faculty of the Graduate School of the University of Kansas. t Present address: Department of Biology and Center for Environmental Studies, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061. 877
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ARTHUR L. BUIKEMA,JR.
active in blue light than in other spectral ranges (Klugh, 1927). Quick shifts from a red to a blue spectrum will provoke a negative phototactic response as will an increase in the light intensity under a given spectral range (Baylor & Smith, 1957). Dark adapted Daphnia pulex and Simocephalus exhibit a temporary decrease in heart rate when they are suddenly exposed to illumination (Schulz, 1928) although this decrease apparently was not observed in D. magna (Proksova, 1950). Exposure of light adapted animals to a decrease in light intensity resulted in a temporary increase in the heart rate (Schulz, 1928). Heart rate of Calanus finmarchicus decreased when light intensity was increased and blue wavelengths were most effective (Harvey, 1930). Schulz’s (1928) data suggest that the effect of light on heart rate is not mediated via the naupliar or compound eye but that light affects the heart itself. The heart rate of the shrimps Palaemonetes (Scudamore, 1941) and Paratya (Hara, 1952) decreased when they were exposed to a dark background and the effect was mediated via the eyes. The foregoing studies all suggest that because light does affect the activity of aquatic organisms there may also be an effect on the rate of oxygen consumption as well. The rate of oxygen consumption of C. jnmarchicus was higher in the light than in the dark (Marshall et al., 1935). However, the presence or absence of light did not significantly affect the oxygen consumption of the copepods C. plumchrus, when tested at various temperatures (Topping, 1965), Acartia claussi and A. tonsa (Conover, 1956) or the euphausiid, Euphausia paci$ca (Pearcy et al., 1969). Berner (1962) suggested that the low metabolic rates obtained for the copepod, Temora Zongicornis, were possibly the result of low activity levels of the organism in the dark. Topping (1965), although he did not demonstrate any long-term effects of light vs. dark on the metabolic rate of Calanus plumchrus, did demonstrate a significant temporary increase in metabolic rate when the copepod was suddenly exposed to illumination, Marshall et al. (1935) reported that beyond a comparatively low light intensity there was no effect of increasing light intensity on metabolic rate. However, they did not state a light intensity value. Studies on the effect of light on metabolic rate are very limited and have been done entirely on marine organisms. The foregoing studies, except for Pearcy et al. (1969), do not report light intensity values. Body size has not been considered as a variable even though the response of organisms to light is known to vary with age (Clarke, 1932; Lumer, 1932; Wechsler & St. John, 1960). Because of the limited nature of the above studies, the following experiment was designed to study the effect of body size, light and light acclimation on the rate of oxygen consumption of the fresh-water cladoceran, D. pulex. MATERIALS AND METHODS Duphniu were obtained from the stock cultures which are maintained at the University of Kansas. Acclimated animals were cultured under their respective light conditions for a minimum of two complete molt cycles and usually for an entire life cycle. Unacclimated animals were selected from a stock tank which was illuminated at about 50 ft-c.
OXYGEN
CONSUMPTION
OF DAPHNIA
PULEX
879
The effects of thirteen light conditions were studied under a 14 hr photoperiod. Eight different light intensities were studied and light intensity was controlled by varying the distance of the cultures from a 40 W cool white deluxe fluorescent bulb (spectral range 350-750 rnp) and the use of Wratten neutral density filters. The light intensities were measured with a YSI Model 65 radiometer and were: darkness (DK, 0), 26, 50, 100, 200, 400, 800 and 1600 ergs/cm8 per set or approximately 0, 1.7, 3.5, 7, 14, 28, 55 and 110 ft-c. Five wavelength ranges were studied and four ranges were controlled with Kodak Wratten Filters. These ranges were: entire visible spectrum (350-750 n-&; 360490 rnp (violet; filter 49B); 460-540 rnp (blue; filter 75); 560-610 q (green; filter 73); and 650-7OO+mp (red; filter 70). The intensities under these wavelength conditions varied from 0 to 1600 ergs/cm* per set for entire spectrum; 11 ergs/cm8 per set for violet, blue and green light; and 32 ergs/cma per set for red light. A B+ L polarizing filter was used for the polarized light condition (POL) and the light intensity was 90 ergs/cm2 per set or about 6.6 ft-c. Throughout this paper the cultures will be referred to by intensity (ft-c), color or other symbol given (DK and POL). The Duphniu were not fed for approximately 12 hr before their rate of oxygen consumption was determined. The animals were tested in pond water which was filtered through a Metricel VM-6 filter with a pore size of 0.45 t.~and aerated for 24 hr prior to use. Glass stoppered bottles, approximately 60 ml capacity, were used as respirometers. The volume of the respirometers was determined with graduated pipettes to within 0.5 ml. At least one control and, depending on the number of available Duphniu, eight to fourteen respirometers were run at each light condition. The number of Duphniu per respirometer varied because of size and abundance. Generally, five adult and twenty-five young were placed in their respective respirometers. The respirometers were stoppered and checked for air bubbles. The respirometers were then placed on their sides in their respective light conditions for 24 hr. At the end of 24 hr the amount of dissolved oxygen in each respirometer was determined by the unmodified Winkler method, using a lo-ml burette calibrated in 0.02 ml. A 0.005 N sodium thiosulfate solution was used. The difference between the amount of oxygen present in a respirometer with animals and the average amount of oxygen in the controls was considered the volume of oxygen consumed per 24 hr by the Duphniu. The volume of oxygen consumed was then corrected to a per animal per hr value. The body length of the Duphniu was measured to the nearest 0.01 mm with an ocular micrometer from the top of the head to the base of the spine. The body weight, in mg, of the Duphniu was then estimated from length by the weight-length relationship of Richman (1958) where Wt = 0.028L - 0.022, (1) where Wt is dry weight in mg and L is length in mm (1). One modification was made in using the above equation (1) in that the minimum weight was considered to be not less than 0.003 mg. Spot checks of the weights of different sized Duphniu used in each phase of this experiment correspond with Richman’s data on the same species (Brooks, 1957) and the use of his equation seems reasonable. Regression coefficients (b value) and an analysis of variance were then calculated for ~1 of oxygen consumed per Duphniu per hr as a function of body weight for each light condition and state of acclimation. The b values and adjusted means obtained for unacclimated and acclimated animals under each light condition were compared statistically (Steele & Torrie, 1960). Additionally all thirteen b values and adjusted means obtained for the unacclimated, and for the acclimated, animals were statistically compared for differences among each other. The b values were first tested for homogeneity and if the b values were not homogeneous, then those b values which were significantly different were determined by using a modified Student-Newman-Keuls procedure (J. Rohlf, personal communication) where the b values were arrayed from smallest to largest and where a significant difference
ARTHUR L. BUIKEMA, JR.
880
between two b values would be equal to or exceed
QS ,@A1 /xX’x~~ + 1 ZXZV.
(2)
The value of Q is found from a table of upper percentage points of the Studentized range and Q varies with the number of b values being compared. The average value of S is derived from all the data where
s=
error sum of squares, N1-2+.
+ . . . + error 7 .+Nj-2
sum of squaresj
(3)
The values of 112 xle and l/ z xaa [equation (2)] are the respective values determined for each of the two b values being compared at any one time. Analyses of covariance were also computed to determine if there were any significant differences among adjusted means (Steele & Torrie, 1960). If the analysis was significant, then those adjusted means which were significantly different from each other were determined by using Finney’s approximation (Steele & Torrie, 1960). When the adjusted means are compared for more than two b values, and because the analysis requires an average b value, certain variations in the analysis were employed. If there were significant differences among b values but no one b value was significantly different from all of the others, an average b value was determined for all of the data (Steele & Torrie, 1960) and used in the analysis of differences among adjusted means. If a b value was significantly different from all the other b values, the data used in the computation of that b value were not used to compute an average b value. Also, Finney’s approximation is easily employed if one is comparing two adjusted means and the data sets from which each adjusted mean is computed are unequal in size. However, if one uses Finney’s approximation to compare many adjusted means and the data sets are of unequal size, one is justified in computing a harmonic value (G. Erickson, personal communication). RESULTS
Light intensity markedly affects the relationship between body size and rate of oxygen consumption of unacclimated Daphniu (F = 19.58, d.f. = 12, 144; PC 0.005; Table 1, Fig. 1). Daphnia tested at l-7 and 3.5 ft-c and in darkness had higher b values than those tested under higher light intensities. The b value for 3.5 ft-c was significantly different from those obtained at all other light intensities. The b value for 1.7 ft-c was significantly different from the b values obtained for 110, 28, 14 and 3.5 ft-c. The b value for dark (DK) was significantly different from those obtained for 110, 14 and 3.5 ft-c, but not from that obtained for 28 ft-c. Interestingly, at intensities above 3.5 ft-c, light intensity did not significantly affect the relationship between body size and rate of oxygen consumption (Table 1; Fig. 1). A comparison among adjusted means (F = 12.33; d.f. = 12, 156; P
OXYGEN
CONSUMPTION
TABLE I-EFFECT ON
THE RELATiONSHIP
OF DAPHNIA
OF LIGHT AND OF OXYGEN
LIGHT
881
PVLEX ACCLIMATION
CONSUMPTION
TO BODY
SIZE
F values b values (S,) t Condition Violet Blue Green Red 110 ft-c 55 ft-c 28 ft-c 14 ft-c 7 ft-c 3.5 ft-c I.7 ft-c DK (0 ft-c) POL (6.6 ft-c)
Ergs/cm* per set 11 11 11 32 1600 800 400 200 100 50 26 0 90
N Unacclimated 14 13 14 13 10 9 9 15 14 15 14 15 15
l-124 0.978 0.908 1‘117 0.674 0.863 0.747 0,714 0.985 I.478 1.203 I.126 0.713
(0.030) (0,043) (0@46) (0.060) (0.057) (0.120) (0.081) (0,048) (0.024) (0.019) (0.032) (O-073) (0~070)
Between b values
N Acclimated 10 10 10 10 9 8 7 9 8 IO 9 10 9
0.770 (0‘087) 0,633 (0.035) 0*380 (0.035) 0838 (O-047) 0839 (0.088) 0.642 (0,074) 0442 (0.086) 1,090 (0.03 5) Oq988 (0,113) 0,799 (0.046) 0.370 (0.073) 0~930 (0*068) 0.836 (0.064)
16.98$ 37.60: 73.28 : 13.22: 2.58 2.26 6*07$ 37-11: 0.00 211.39: 98-13: 3.38 1.36
Between adjusted means
--T 27.48 : 0.07
9*26* s-74: 4494:
i AI1 values of b are significantly different from zero, P < O-005. 1 F value is significant, P < 0*005 except for (*) where 0.005
FIG. 1. The relationship between oxygen consumption and light intensity for unacclimated (0) and acclimated (0) Z&@&r of different sizes (0.7 mm = 0.003 mg; 1.4 mm = 0.017 mg; 2.1 mm = 0.037 mg; and 2.8 mm = 0.056 mg). The affect of polarized light (POL; 6.6 ft-c) on the oxygen consumption of unacclimated (A) and acclimated (0) animals is also indicated, 3’
882
ARTHUR L. BUIKEMA,JR.
above 28 ft-c. The effects of 1.7 and 3.5 ft-c are attributed to the significant differences among b values. Acclimation of Duphniu to different light conditions generally resulted in a lower b value (Table 1). The b values obtained for acclimated 1.7, 3.5 and 28 ft-c were significantly lower than those obtained for unacclimated animals and the b value for 14 ft-c was significantly higher. Although there were no significant differences between the b values for unacclimated and acclimated animals tested under DK, 7 and 110 ft-c, there was a significant difference between adjusted means (Table 1). The adjusted means for acclimated DK and 110 were significantly larger than those for unacclimated animals, whereas the adjusted mean for acclimated 7 was significantly lower than that for unacclimated animals (Table 1). An analysis for homogeneity of acclimated b values was significant (F = 10.47; d.f. = 12, 93; P-e 0.005). The b value obtained for 14 ft-c was significantly larger than those obtained for 1.7, 28 and 55 ft-c. In addition, the b values for 1.7 and 28 ft-c were significantly smaller than that obtained for 7 ft-c. There were fewer significant differences among the b values obtained for acclimated animals and those b values which were significantly different among unacclimated animals were not significantly different after acclimation. There were significant differences among adjusted means for acclimated animals (F = 13.45; d.f. = 12, 105; P-CO-005). The adjusted mean for 110 ft-c was significantly larger than that of any other light condition. The adjusted mean for 55 ft-c was significantly higher than DK, 1.7 and 3.5 ft-c; 28 ft-c was significantly larger than DK, 1.7 and 3.5 ft-c; and 1.7 ft-c was also smaller than 3.5 ft-c and DK. These results also suggest that acclimation affects the position of the regression slopes in a manner different from those compared for unacclimated animals. The effect of light acclimation and intensity on the metabolism of differentsized Duphniu suggests that the stimulatory effect of 1.7 and 28 ft-c and the depressant effect of 3.5, 7 and 14 ft-c on small animals reverses as the animals become larger in size and the effects become respectively depressant and stimulatory. Above 14 ft-c the results suggest that although there is little effect of light intensity on the metabolism of small Duphniu, the rate of oxygen consumption linearly increases with light intensity as the Duphniu become larger (Fig. 1). Furthermore, acclimation to light does affect the metabolism of different-sized Duphniu and the effect varies with size and light intensity. The effect of 1.7 ft-c is probably due to a combination of a small b value and significantly smaller adjusted mean. The effects of 3.5, 7 and 14 ft-c are in part due to differences in adjusted means while the effects of intensities above 14 ft-c are due to relatively high adjusted means. Polarized light (6.6 ft-c) did not significantly affect the b values of acclimated and unacclimated animals (Table 1) and the b values were not significantly different from those obtained at 7 ft-c. Consequently, the effect of polarized light on the relationship between metabolic rate and body size is probably inseparable from
OXYGEN
CONSUMPTION
OF DAPHNIA
883
PULEX
the effect of light intensity. However, because the adjusted means of acclimated animals between polarized light and 7 ft-c are significantly different, polarized light affects metabolic rate. Polarized light depresses metabolism of unacclimated animals and stimulates that of acclimated animals when compared with each other (Table 1; Fig. 1) and when compared to 7 ft-c. The rate of oxygen consumption of animals acclimated to polarized light is significantly higher than that of unacclimated animals (Table 1) at all sizes of D@niu (Fig. 1). The b values were significantly lower for animals acclimated to the wavelength ranges than for unacclimated animals (Table 1). The b values for unacclimated and acclimated animals were homogeneous in both cases, and there were no significant differences among adjusted means. Wavelength essentially had no effect on animals weighing 0.017 mg or larger although red wavelengths may have had a slightly depressant effect on the metabolic rate of young animals while green wavelengths may have been slightly stimulatory (Fig. 2). Except for small l.OOt .60 A0 -520 . ._0 +10: 1 e.06 0104 2 9 .02 -
A/oo3mg
.OlC ’ VIOLET
I
I
I
BLUE
GREEN
RED
FIG. 2. The relationship between oxygen consumption and wavelength for unacclimated (0) and acclimated (0) Daphniu of different sizes (0.7 mm = 0.003 mg; 1.4 mm = 0,017 mg; 2.1 mm = O-037 mg; and 2.8 mm = 0.056 mg). animals
(0,003 mg) the effects of acclimation to wavelength resulted in a lower rate of oxygen consumption. Because the light intensity under the violet, blue and green filters is the same (11 ergs/cm2 per set) and because there were no significant differences among b values and adjusted means, there is no independent effect of wavelength on metabolic rate that is separable from the effect of light intensity. Because the intensity under the red filter (32 ergs/cm2 per set) is close to 1.7 ft-c (26 ergs/cm2 per set) there is probably no independent effect on the b values of unacclimated and
884
ARTHUR L. BUIKEMA, JR.
acclimated animals nor is there a significant difference between adjusted means of unacclimated animals. However, acclimation to 1.7 ft-c did result in a significantly lower adjusted mean than that obtained for red wavelengths suggesting that a drop in average metabolic rate may be due to acclimation to a broad spectral range rather than to intensity or red wavelengths per se. DISCUSSION
Although the metabolism of crustaceans as a function of various environmental parameters has been studied frequently (Richman, 1958; Wolvekamp & Waterman, 1960) relatively few studies were concerned with the effect of these parameters on the relationship between rate of oxygen consumption and body size. Some workers have expressed their data as a function of body length (Obreshkove, 1930; Jancarik, 1948) while others have expressed their data relative to the metabolic rate of the first instar (Terao, 1931; Hoshi, 1949). The latter studies are not easily compared with other studies. The relationship between rate of oxygen consumption and body weight is described by the equation vol. 0, = aI&%,
(4)
where W is dry weight, b is the regression coefficient and a is the intercept value. The b value for crustacea generally varies between 0.67 and 1.00 (Wolvekamp & Waterman, 1960). A b value of 0.67 is interpreted to denote a relationship between metabolism and body surface area while a value of 1.00 has been described as a relationship between metabolism and body weight. Intermediate values, between 0.67 and l-00, are found for the majority of crustaceans. Weymouth et al. (1944) reported a mean b value of 0826 for a number of species of crustaceans and Zeuthen (1953) reported that the b value for organisms less than 1 mg nitrogen is O-95. Richman (1958) reported a b value of 0.881 for D. pulex. The b values obtained in this study are in general agreement when one compares the average b value determined for unacclimated animals, O-930 (0.979 if the data for 3.5 ft-c are included), and acclimated animals, O-780, although there is considerable variation among b values (Table 1). Interestingly, the b values are generally higher for unacclimated animals. Variation among b values is not unusual and they vary with temperature for snails (Berg & Ockelman, 1959) and amphipods (Armitage, 1962) and among various genera and/or species of marine copepods (Conover, 1959), crabs (Teal, 1959) and other crustaceans (Wolvekamp & Waterman, 1960). The results of this study suggest that light also influences the relationship between metabolism and body weight and that this relationship is also affected by light acclimation. The effect of light acclimation is more evident if the data obtained for the animals in the dark are compared to Richman’s data for the same species since Richman (1958) also tested his animals in the dark. The b value obtained by Richman (calculated from his data, 1958) is sigmficantly different from the b value obtained for unacclimated animals (F = 22.98, PC 0*005). Because the animals
OXYGEN
CONSUMPTION
OF
DAPHNIA
PVLEX
885
used in this study were either not acclimated or acclimated for at least two complete molt cycles, the significant differences between my data and Richman’s are difficult to interpret. The significant difference in the b values may be an artifact, or it may be due to the fact that Richman’s animals were partially acclimated to the dark. Richman states that the animals were placed in filtered water 24 hr prior to the experiment but he does not state whether they were kept in the dark. He did adapt his animals to the dark in his feeding study. Because the adjusted means between animals not acclimated and acclimated to the dark are significantly different (this study) and also the adjusted mean between Richman’s data and that for acclimated animals is significantly different, it is possible that acclimation to the dark may be a type of translation. However, the significant difference between Richman’s data and mine for dark acclimated animals may be due to the fact that Richman’s animals were more crowded (fifty Da@& per 5 ml and 50-100/135 ml vs. five to twenty-five Duphniu per 60 ml) when he determined oxygen consumption. Crowding resulted in a lower rate of oxygen consumption for 5’. vet&s (Hoshi, 1957) but had an opposite effect on D. magna (Zeiss, 1963). If crowding caused an increase in oxygen consumption of D. pulex, I would have expected Richman’s results to be the same or higher than mine and they were not. If the data obtained in this experiment are expressed as ~1. O,/mg per hr, they compare well with those obtained by other authors as summarized by Richman (1958). As Richman noted, the weight specific rate of oxygen consumption of D. magna, Chydorus ovalis and C. Jinmarchicus are not in agreement and may be due to their larger size. Because light intensity is known to affect the activity of aquatic organisms (Yerkes, 1900; Dice, 1914; Schulz, 1928; Clarke, 1932; Ringelberg, 1963, 1964) one would expect that a change in metabolism should be demonstrable. Interestingly, Marshall et al. (1935) reported that the activity of C. jinmarchicus did not change when it was exposed to light even though the rate of oxygen consumption increased over 100 per cent. Activity was not observed in this experiment; but, because the swimming velocity of Duphniu increases as light intensity decreases, one might expect that metabolic rate would be higher at low light intensities. This was not necessarily the case (Fig. 1). Interestingly, for unacclimated animals there is no significant effect of light intensities above 14 ft-c on metabolic rate (Fig. 1). Marshall et al. (1935) reported an increased metabolic rate for C. Jinmarchicus when light intensity increased and that under subsequent increases in light intensity there was no effect, In acclimated organisms there is an increase in the rate of oxygen consumption above 14 ft-c. While the light intensities studied in this experiment are not very high, there is an effect of light increasing the metabolic rate of organisms, but the interpretation of the results depends on the light intensities being compared, acclimation state and size of the animals. In those organisms where there was no significant effect of light or dark on metabolic rate (Conover, 1956; Topping, 1965 ; Pearcy et al., 1969) the light intensities which were used may have
886
ARTHUR
L.
BUIKEMA,
JR.
coincidentally resulted in similar data and had other light intensities been used, the results may have been different. For example, the rates of oxygen consumption obtained for unacclimated, large animals (0.056 mg) in the dark, 28,55 and 110 ft-c are probably not significantly different even though the rate obtained for 14 ft-c is significantly lower (Fig. 1). Baylor & Smith (1953, 1957) suggest that the response of Daphnia to polarized light is due to an increase in the intensity of light which enters the cone lenses of the eye. The effect of polarized light (66 ft-c) on the b value was inseparable from the affect of 7 ft-c, but the adjusted means obtained for animals tested under polarized light are significantly different from that obtained at 7 ft-c but not from 14 ft-c (unacclimated) or 28 and 55 ft-c (acclimated) which suggests that the effect of polarized light on the rate of oxygen consumption may be due to an apparent higher intensity perceived by the animal. Previously cited studies suggest that wavelengths, because they affect activity, may also affect the rate of oxygen consumption. The results obtained in this experiment do not support such a hypothesis (Fig. 2). Even though there is a considerable difference in the nature and velocity of the blue dance, which is associated with food-hunting, and the red dance, which maintained the organism in an area of high food concentration (Baylor & Smith, 1957; Smith & Baylor, 1959) it is suggested that a Daphnia which is maintaining its position in an area of high food concentration expends as much energy as a Daphnia searching for food. SUMMARY 1.
The effects of nine light intensities (0, 1.7, 3.5, 7, 14, 28, 55 and 110 ft-c), polarized light (6.6 ft-c) and four wavelength ranges were studied on the relationship (b) of metabolic rate and body size for unacclimated and acclimated Daphnia pulex. 2. Light intensity significantly affects b and the effect varies with acclimation. Light intensities of 1.7 and 3.5 ft-c were stimulatory, 14 ft-c was depressant and intensities above 28 ft-c were slightly depressant on the metabolic rate of different sized unacclimated animals. Light intensities of 1.7 and 28 ft-c were stimulatory and intensities of 3.5, 7 and 14 ft-c were depressant on the metabolic rate of young animals and the effects were reversed in older animals. Above 14 ft-c the metabolic rate of older acclimated animals increases linearly with light intensity. 3. The effect of polarized light on b was inseparable from the effect of light intensity but there were significant differences among adjusted means. Acclimation to polarized light resulted in a significantly higher adjusted mean. 4. There was no significant effect of wavelength among b values, and adjusted mean values, for unacclimated and acclimated animals. Acclimation to each wavelength did result in a significantly lower b value. Acknowledgements-I encouragement
am indebted to my major and co-operation during this study.
advisor,
Dr.
K.
B. Armitage,
for his
OXYGEN
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OF DAPHNIA
PVLEX
887
REFERENCES ARMITAGEK. B. (1962) Temperature and oxygen consumption of Orchomonella chilensis. Biol. Bull. mar. biol. Lab., Woods Hole 123, 225-232. BAINBRIDGER. & WATERMANT. H. (1958) Turbidity and the polarized light orientation of the crustacean Mysidium. J. exp. Biol. 35, 487-493. BAYLORE. R. & SMITH F. E. (1953) The orientation of Cladocera to polarized light. Am. Nat. 87, 97-101. BAYLORE. R. & SMITH F. E. (1957) Diurnal migration of plankton crustaceans. In Recent Advances in Invertebrate Physiology (Edited by SCHEERB. T.), pp. 21-35. University of Oregon, Oregon. BERG K. & OCKELMANK. W. (1959) The respiration of freshwater snails. J. exp. Biol. 36, 690-708. BERNER A. (1962) Feeding and respiration in the copepod, Temora longicornis (Muller). J. mar. Biol. Ass. U.K. 42, 625-640. BROOKSJ. L. (1957) The systematics of North American Daphnia. Mem. Corm. Acad. Arts and Sci. 13, I-180. CLARKEG. L. (1930) Change of phototropic and geotropic signs in Daphnia induced by changes in light intensity. J. exp. Biol. 7, 109-131. CLARKEG. L. (1932) Quantitative aspects of the change of phototrophic sign in Daphnia. J. exp. Biol. 9, 180-211. CONOVERR. J. (1956) Oceanography of Long Island Sound-VI. Biology of Acartia clausi and A. tonsa. Bull. Bingham Oceanogr. Collect. Yale Univ. 15, 156-233. CONOVERR. J. (1959) Regional and seasonal variation in the respiratory rate of marine copepods. Limnol. Oceanogr. 4, 259-268. GUSHINGD. H. (1951) Some experiments on the vertical migration of zooplankton. r. Anim. Ecol. 24, 137-166. DICE L. R. (1914) The factors determining the vertical movements of Daphnia. J. Anim. Behav. 4, 229-236. HARAJ. (1952) On the hormones regulating the frequency of the heart beat in the shrimp, Paratya compressa. Annot. 2001. Japon. 25, 162-171. HARVEYJ. M. (1930) The action of light on Calanus finmarchicus (Gunner) as determined by its effect on the heart rate. Contr. Can. Biol. Fish, 5, 85-92. HAZEN W. E. & BAYLORE. R. (1962) Behavior of Daphnia in polarized light. Biol. Bull. 123, 243-252. HOSHI T. (1949) Studies on physiology and ecology of plankton-II. Oxygen consumption and heart-beat rate of developmental stages of Simocephalus vetulus (0. F. Muller). Sci. Rep. Tohoku Univ. Ser. 4. 18, 159-161. HOSHI T. (1957) Studies on physiology and ecology of plankton-XII. Changes in 0, consumption of the daphnid, Simocephalus vetulus, with the decrease of Or-concentration. Sci. Rep. Tohoku Univ. Ser. 5, 33, 27-33. JANCARIKA. (1948) Contribution to the knowledge of the physiology of breathing of cladocera, Daphniapulex. Publ. Fat. Sci. Univ. Masaryk, Rada M 1.305, l-42. KLUGH A. B. (1927) The ecology, food relations and culture of freshwater entomostraca. Roy. Can. Inst., Trans. 16, 15-98. LUMER H. (1932) The reactions of certain Cladocera to colored lights of equal intensity. OhioJ. Sci. 32, 218-231. MARSHALLS. M., NICHOLLSA. G. & ORR A. P. (1935) On the biology of Calanus jinmarchicus-VI. Oxygen consumption in relation to environmental conditions. r. mar. Biol. Ass. U.K. 37, 459-472. NAGANOT. (1955) The behavior of crustacea exposed to colored light. Bull. Mar. Biol. Sta. Asamushi 7, 44-49.
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OBRESHKOVE V. (1930) Oxygen consumption in the developmental stages of a cladoceran. Physiol. 2001. 3, 271-282. PEARCYW. G., THEILACKER G. H. & LASKERR. (1969) Oxygen consumption of Euphausiu pacifica : the lack of a die1 rhythm or light-dark effect, with a comparison of experimental techniques. Limnol. Oceanogr. 14, 219-223. PROK~OVA M. (1950) 0 srde&ii Einnosti perloocky Duphniu magna Strauss. Acta Acud. Sci. Nat. Moruv. 22, 33-56. RICHMAN S. (1958) The transformation of energy by Daphniu pulex. Ecol. Monogr. 28, 273-291. RINGELBERC J. (1963) The behavior of Daphniu in diffuse light. Nuturwiss. 50,313-314. RINGELBERG J. (1964) The positively phototactic reaction of Duphniu magna Straus. Neth.J. Sea Res. 2, 319-406. SCHULZ H. (1928) Uber die Bedeutung des Lichtes im Leben neiderer Krebse. Z. vergl. Physiol. 7, 488-552. SCUDAMORE H. H. (1941) A correlation between rate of heart beat and the state of certain chromatophores in the shrimp Pulaemonetes. Illinois State Acud. Sci., Trans. 34,238-240. SMITH F. E. & BAYLORE. R. (1953) Color responses in the Cladocera and their ecological significance. Am. Nut. 57, 49-55. STEELER. G. D. & TORRIEJ. H. (1960) Principles and Procedures of Statistics. McGraw-Hill, New York. TEAL J. M. (1959) Respiration of crabs in Georgia salt marshes and its relation to their ecology. Physiol. Zoiil. 32, I-14. TERAO A. (1931) Change in vitality with age based on the living unit of organisms-I. Oxygen consumption in the daphnid, Simocephulus expinosns. Jup. Acud., Tokyo, Proc. I, 23-25. TOPPING M. S. (1965) A study of oxygen consumption in Culanus plumchrus Marukawa 1921 and implications on vertical migration. Master’s thesis, University of British Columbia. WECHSLER A. S. & ST. JOHNP. A. (1960) The spectral sensitivity of Daphniu magna and its variance with age. Anat. Rec. 137, 400 (Abstr.). WEYMOUTHF. W., CRISMONJ. M., HALL V. E., BELDINGH. S. & FIELDJ., II (1944) Total and tissue respiration in relation to body weight. Physiol. Zodl. 17, 50-71. WOLVEKAMPH. P. & WATERMANT. H. (1960) Respiration. In Physiology of Crustuceu. Metabolism and Growth (Edited by WATERMANT. H.), Vol. I, pp. 35-100. Academic Press, New York. YERKESR. M. (1900) Reaction of Entomostraca to stimulation by light-II. Reactions of Daphniu and Cypris. Am. J. Physiol. 4, 405-422. ZEISS F. E., JR. (1963) Effects of population densities on zooplankton respiration rates. Limnol. Oceunogr. 8, 110-115. ZEUTHENE. (1953) Oxygen uptake as related to body size in organisms. Q. Rev. Biol. 28, l-12. Key Word Index-Oxygen body size.
consumption ; cladocera ; Daphnia pulex ; light ; acclimation ;
OXYGEN CONSUMPTION OF THE CLADOCERAN, DAPHNIA PULEX, AS A FUNCTION OF BODY SIZE, LIGHT AND LIGHT ACCLIMATION* ARTHUR
L.
BUIKEMA,
JR.~
Department of Zoology, University of Kansas, Lawrence, Kansas 66044
(Received 15 November 1971)
The effects of nine light intensities, polarized light and four wavelength ranges were studied on the metabolic rate of Duphnia. 2. The effect of light intensity was significant and the effects varied depending on, light intensity, body size and state of acclimation. 3. Except for differences among adjusted means the effect of polarized light was inseparable from the effect of light intensity. 4. Acclimation to wavelength reduced metabolic rate and there were no other significant effects.
Abstract-l.
INTRODUCTION
LIGHT intensity affects vertical migration and changes in locomotory activity of Cladocera (Yerkes, 1900; Dice, 1914; Schulz, 1928; Clarke, 1930, 1932; Cushing, 1951; Berner, 1962; Ringelberg, 1963, 1964) and the response to light intensity varies with age (Clarke, 1932). Polarized light also affects the behavior of aquatic organisms and the effect has been described as one of increasing light intensity in the cone lenses (Baylor & Smith, 1953, 1957; Bainbridge & Waterman, 1958; Hazen & Baylor, 1962). Wavelength also affects activity and red wavelengths (620-640 rnp) are most effective in stimulating the activity of Daphnia pulex, D. magna, Leptodora kindtii and Moina. Lesser stimulatory wavelengths occurred around 540 rnp for Moina only and 440 rnp for the other species (Lumer, 1932). Blue wavelengths (400450 rnp) may be more stimulatory than red wavelengths (680-700 rnp) for Simocephalus vetulus and the shrimp, Paratya compressa, but there are no data for the 550-680 rnp range (Nagano, 1955). Smith & Baylor (1953) and Baylor & Smith (1957) report that blue and red dances are exhibited by cladocerans, copepods, fairy shrimp and SquiZZu larvae and that blue wavelengths distinctly agitate the populations so that the velocity of the blue dance is three to five times that exhibited in the red dance. The ostracod, Cyprinotus incongruens, was more * This paper is part of a Ph.D. thesis submitted to the Faculty of the Graduate School of the University of Kansas. t Present address: Department of Biology and Center for Environmental Studies, Virginia Polytechnic Institute and State University, Blacksburg, Virginia 24061. 877
878
ARTHUR L. BUIKEMA,JR.
active in blue light than in other spectral ranges (Klugh, 1927). Quick shifts from a red to a blue spectrum will provoke a negative phototactic response as will an increase in the light intensity under a given spectral range (Baylor & Smith, 1957). Dark adapted Daphnia pulex and Simocephalus exhibit a temporary decrease in heart rate when they are suddenly exposed to illumination (Schulz, 1928) although this decrease apparently was not observed in D. magna (Proksova, 1950). Exposure of light adapted animals to a decrease in light intensity resulted in a temporary increase in the heart rate (Schulz, 1928). Heart rate of Calanus finmarchicus decreased when light intensity was increased and blue wavelengths were most effective (Harvey, 1930). Schulz’s (1928) data suggest that the effect of light on heart rate is not mediated via the naupliar or compound eye but that light affects the heart itself. The heart rate of the shrimps Palaemonetes (Scudamore, 1941) and Paratya (Hara, 1952) decreased when they were exposed to a dark background and the effect was mediated via the eyes. The foregoing studies all suggest that because light does affect the activity of aquatic organisms there may also be an effect on the rate of oxygen consumption as well. The rate of oxygen consumption of C. jnmarchicus was higher in the light than in the dark (Marshall et al., 1935). However, the presence or absence of light did not significantly affect the oxygen consumption of the copepods C. plumchrus, when tested at various temperatures (Topping, 1965), Acartia claussi and A. tonsa (Conover, 1956) or the euphausiid, Euphausia paci$ca (Pearcy et al., 1969). Berner (1962) suggested that the low metabolic rates obtained for the copepod, Temora Zongicornis, were possibly the result of low activity levels of the organism in the dark. Topping (1965), although he did not demonstrate any long-term effects of light vs. dark on the metabolic rate of Calanus plumchrus, did demonstrate a significant temporary increase in metabolic rate when the copepod was suddenly exposed to illumination, Marshall et al. (1935) reported that beyond a comparatively low light intensity there was no effect of increasing light intensity on metabolic rate. However, they did not state a light intensity value. Studies on the effect of light on metabolic rate are very limited and have been done entirely on marine organisms. The foregoing studies, except for Pearcy et al. (1969), do not report light intensity values. Body size has not been considered as a variable even though the response of organisms to light is known to vary with age (Clarke, 1932; Lumer, 1932; Wechsler & St. John, 1960). Because of the limited nature of the above studies, the following experiment was designed to study the effect of body size, light and light acclimation on the rate of oxygen consumption of the fresh-water cladoceran, D. pulex. MATERIALS AND METHODS Duphniu were obtained from the stock cultures which are maintained at the University of Kansas. Acclimated animals were cultured under their respective light conditions for a minimum of two complete molt cycles and usually for an entire life cycle. Unacclimated animals were selected from a stock tank which was illuminated at about 50 ft-c.
OXYGEN
CONSUMPTION
OF DAPHNIA
PULEX
879
The effects of thirteen light conditions were studied under a 14 hr photoperiod. Eight different light intensities were studied and light intensity was controlled by varying the distance of the cultures from a 40 W cool white deluxe fluorescent bulb (spectral range 350-750 rnp) and the use of Wratten neutral density filters. The light intensities were measured with a YSI Model 65 radiometer and were: darkness (DK, 0), 26, 50, 100, 200, 400, 800 and 1600 ergs/cm8 per set or approximately 0, 1.7, 3.5, 7, 14, 28, 55 and 110 ft-c. Five wavelength ranges were studied and four ranges were controlled with Kodak Wratten Filters. These ranges were: entire visible spectrum (350-750 n-&; 360490 rnp (violet; filter 49B); 460-540 rnp (blue; filter 75); 560-610 q (green; filter 73); and 650-7OO+mp (red; filter 70). The intensities under these wavelength conditions varied from 0 to 1600 ergs/cm* per set for entire spectrum; 11 ergs/cm8 per set for violet, blue and green light; and 32 ergs/cma per set for red light. A B+ L polarizing filter was used for the polarized light condition (POL) and the light intensity was 90 ergs/cm2 per set or about 6.6 ft-c. Throughout this paper the cultures will be referred to by intensity (ft-c), color or other symbol given (DK and POL). The Duphniu were not fed for approximately 12 hr before their rate of oxygen consumption was determined. The animals were tested in pond water which was filtered through a Metricel VM-6 filter with a pore size of 0.45 t.~and aerated for 24 hr prior to use. Glass stoppered bottles, approximately 60 ml capacity, were used as respirometers. The volume of the respirometers was determined with graduated pipettes to within 0.5 ml. At least one control and, depending on the number of available Duphniu, eight to fourteen respirometers were run at each light condition. The number of Duphniu per respirometer varied because of size and abundance. Generally, five adult and twenty-five young were placed in their respective respirometers. The respirometers were stoppered and checked for air bubbles. The respirometers were then placed on their sides in their respective light conditions for 24 hr. At the end of 24 hr the amount of dissolved oxygen in each respirometer was determined by the unmodified Winkler method, using a lo-ml burette calibrated in 0.02 ml. A 0.005 N sodium thiosulfate solution was used. The difference between the amount of oxygen present in a respirometer with animals and the average amount of oxygen in the controls was considered the volume of oxygen consumed per 24 hr by the Duphniu. The volume of oxygen consumed was then corrected to a per animal per hr value. The body length of the Duphniu was measured to the nearest 0.01 mm with an ocular micrometer from the top of the head to the base of the spine. The body weight, in mg, of the Duphniu was then estimated from length by the weight-length relationship of Richman (1958) where Wt = 0.028L - 0.022, (1) where Wt is dry weight in mg and L is length in mm (1). One modification was made in using the above equation (1) in that the minimum weight was considered to be not less than 0.003 mg. Spot checks of the weights of different sized Duphniu used in each phase of this experiment correspond with Richman’s data on the same species (Brooks, 1957) and the use of his equation seems reasonable. Regression coefficients (b value) and an analysis of variance were then calculated for ~1 of oxygen consumed per Duphniu per hr as a function of body weight for each light condition and state of acclimation. The b values and adjusted means obtained for unacclimated and acclimated animals under each light condition were compared statistically (Steele & Torrie, 1960). Additionally all thirteen b values and adjusted means obtained for the unacclimated, and for the acclimated, animals were statistically compared for differences among each other. The b values were first tested for homogeneity and if the b values were not homogeneous, then those b values which were significantly different were determined by using a modified Student-Newman-Keuls procedure (J. Rohlf, personal communication) where the b values were arrayed from smallest to largest and where a significant difference
ARTHUR L. BUIKEMA, JR.
880
between two b values would be equal to or exceed
QS ,@A1 /xX’x~~ + 1 ZXZV.
(2)
The value of Q is found from a table of upper percentage points of the Studentized range and Q varies with the number of b values being compared. The average value of S is derived from all the data where
s=
error sum of squares, N1-2+.
+ . . . + error 7 .+Nj-2
sum of squaresj
(3)
The values of 112 xle and l/ z xaa [equation (2)] are the respective values determined for each of the two b values being compared at any one time. Analyses of covariance were also computed to determine if there were any significant differences among adjusted means (Steele & Torrie, 1960). If the analysis was significant, then those adjusted means which were significantly different from each other were determined by using Finney’s approximation (Steele & Torrie, 1960). When the adjusted means are compared for more than two b values, and because the analysis requires an average b value, certain variations in the analysis were employed. If there were significant differences among b values but no one b value was significantly different from all of the others, an average b value was determined for all of the data (Steele & Torrie, 1960) and used in the analysis of differences among adjusted means. If a b value was significantly different from all the other b values, the data used in the computation of that b value were not used to compute an average b value. Also, Finney’s approximation is easily employed if one is comparing two adjusted means and the data sets from which each adjusted mean is computed are unequal in size. However, if one uses Finney’s approximation to compare many adjusted means and the data sets are of unequal size, one is justified in computing a harmonic value (G. Erickson, personal communication). RESULTS
Light intensity markedly affects the relationship between body size and rate of oxygen consumption of unacclimated Daphniu (F = 19.58, d.f. = 12, 144; PC 0.005; Table 1, Fig. 1). Daphnia tested at l-7 and 3.5 ft-c and in darkness had higher b values than those tested under higher light intensities. The b value for 3.5 ft-c was significantly different from those obtained at all other light intensities. The b value for 1.7 ft-c was significantly different from the b values obtained for 110, 28, 14 and 3.5 ft-c. The b value for dark (DK) was significantly different from those obtained for 110, 14 and 3.5 ft-c, but not from that obtained for 28 ft-c. Interestingly, at intensities above 3.5 ft-c, light intensity did not significantly affect the relationship between body size and rate of oxygen consumption (Table 1; Fig. 1). A comparison among adjusted means (F = 12.33; d.f. = 12, 156; P
OXYGEN
CONSUMPTION
TABLE I-EFFECT ON
THE RELATiONSHIP
OF DAPHNIA
OF LIGHT AND OF OXYGEN
LIGHT
881
PVLEX ACCLIMATION
CONSUMPTION
TO BODY
SIZE
F values b values (S,) t Condition Violet Blue Green Red 110 ft-c 55 ft-c 28 ft-c 14 ft-c 7 ft-c 3.5 ft-c I.7 ft-c DK (0 ft-c) POL (6.6 ft-c)
Ergs/cm* per set 11 11 11 32 1600 800 400 200 100 50 26 0 90
N Unacclimated 14 13 14 13 10 9 9 15 14 15 14 15 15
l-124 0.978 0.908 1‘117 0.674 0.863 0.747 0,714 0.985 I.478 1.203 I.126 0.713
(0.030) (0,043) (0@46) (0.060) (0.057) (0.120) (0.081) (0,048) (0.024) (0.019) (0.032) (O-073) (0~070)
Between b values
N Acclimated 10 10 10 10 9 8 7 9 8 IO 9 10 9
0.770 (0‘087) 0,633 (0.035) 0*380 (0.035) 0838 (O-047) 0839 (0.088) 0.642 (0,074) 0442 (0.086) 1,090 (0.03 5) Oq988 (0,113) 0,799 (0.046) 0.370 (0.073) 0~930 (0*068) 0.836 (0.064)
16.98$ 37.60: 73.28 : 13.22: 2.58 2.26 6*07$ 37-11: 0.00 211.39: 98-13: 3.38 1.36
Between adjusted means
--T 27.48 : 0.07
9*26* s-74: 4494:
i AI1 values of b are significantly different from zero, P < O-005. 1 F value is significant, P < 0*005 except for (*) where 0.005
FIG. 1. The relationship between oxygen consumption and light intensity for unacclimated (0) and acclimated (0) Z&@&r of different sizes (0.7 mm = 0.003 mg; 1.4 mm = 0.017 mg; 2.1 mm = 0.037 mg; and 2.8 mm = 0.056 mg). The affect of polarized light (POL; 6.6 ft-c) on the oxygen consumption of unacclimated (A) and acclimated (0) animals is also indicated, 3’
882
ARTHUR L. BUIKEMA,JR.
above 28 ft-c. The effects of 1.7 and 3.5 ft-c are attributed to the significant differences among b values. Acclimation of Duphniu to different light conditions generally resulted in a lower b value (Table 1). The b values obtained for acclimated 1.7, 3.5 and 28 ft-c were significantly lower than those obtained for unacclimated animals and the b value for 14 ft-c was significantly higher. Although there were no significant differences between the b values for unacclimated and acclimated animals tested under DK, 7 and 110 ft-c, there was a significant difference between adjusted means (Table 1). The adjusted means for acclimated DK and 110 were significantly larger than those for unacclimated animals, whereas the adjusted mean for acclimated 7 was significantly lower than that for unacclimated animals (Table 1). An analysis for homogeneity of acclimated b values was significant (F = 10.47; d.f. = 12, 93; P-e 0.005). The b value obtained for 14 ft-c was significantly larger than those obtained for 1.7, 28 and 55 ft-c. In addition, the b values for 1.7 and 28 ft-c were significantly smaller than that obtained for 7 ft-c. There were fewer significant differences among the b values obtained for acclimated animals and those b values which were significantly different among unacclimated animals were not significantly different after acclimation. There were significant differences among adjusted means for acclimated animals (F = 13.45; d.f. = 12, 105; P-CO-005). The adjusted mean for 110 ft-c was significantly larger than that of any other light condition. The adjusted mean for 55 ft-c was significantly higher than DK, 1.7 and 3.5 ft-c; 28 ft-c was significantly larger than DK, 1.7 and 3.5 ft-c; and 1.7 ft-c was also smaller than 3.5 ft-c and DK. These results also suggest that acclimation affects the position of the regression slopes in a manner different from those compared for unacclimated animals. The effect of light acclimation and intensity on the metabolism of differentsized Duphniu suggests that the stimulatory effect of 1.7 and 28 ft-c and the depressant effect of 3.5, 7 and 14 ft-c on small animals reverses as the animals become larger in size and the effects become respectively depressant and stimulatory. Above 14 ft-c the results suggest that although there is little effect of light intensity on the metabolism of small Duphniu, the rate of oxygen consumption linearly increases with light intensity as the Duphniu become larger (Fig. 1). Furthermore, acclimation to light does affect the metabolism of different-sized Duphniu and the effect varies with size and light intensity. The effect of 1.7 ft-c is probably due to a combination of a small b value and significantly smaller adjusted mean. The effects of 3.5, 7 and 14 ft-c are in part due to differences in adjusted means while the effects of intensities above 14 ft-c are due to relatively high adjusted means. Polarized light (6.6 ft-c) did not significantly affect the b values of acclimated and unacclimated animals (Table 1) and the b values were not significantly different from those obtained at 7 ft-c. Consequently, the effect of polarized light on the relationship between metabolic rate and body size is probably inseparable from
OXYGEN
CONSUMPTION
OF DAPHNIA
883
PULEX
the effect of light intensity. However, because the adjusted means of acclimated animals between polarized light and 7 ft-c are significantly different, polarized light affects metabolic rate. Polarized light depresses metabolism of unacclimated animals and stimulates that of acclimated animals when compared with each other (Table 1; Fig. 1) and when compared to 7 ft-c. The rate of oxygen consumption of animals acclimated to polarized light is significantly higher than that of unacclimated animals (Table 1) at all sizes of D@niu (Fig. 1). The b values were significantly lower for animals acclimated to the wavelength ranges than for unacclimated animals (Table 1). The b values for unacclimated and acclimated animals were homogeneous in both cases, and there were no significant differences among adjusted means. Wavelength essentially had no effect on animals weighing 0.017 mg or larger although red wavelengths may have had a slightly depressant effect on the metabolic rate of young animals while green wavelengths may have been slightly stimulatory (Fig. 2). Except for small l.OOt .60 A0 -520 . ._0 +10: 1 e.06 0104 2 9 .02 -
A/oo3mg
.OlC ’ VIOLET
I
I
I
BLUE
GREEN
RED
FIG. 2. The relationship between oxygen consumption and wavelength for unacclimated (0) and acclimated (0) Daphniu of different sizes (0.7 mm = 0.003 mg; 1.4 mm = 0,017 mg; 2.1 mm = O-037 mg; and 2.8 mm = 0.056 mg). animals
(0,003 mg) the effects of acclimation to wavelength resulted in a lower rate of oxygen consumption. Because the light intensity under the violet, blue and green filters is the same (11 ergs/cm2 per set) and because there were no significant differences among b values and adjusted means, there is no independent effect of wavelength on metabolic rate that is separable from the effect of light intensity. Because the intensity under the red filter (32 ergs/cm2 per set) is close to 1.7 ft-c (26 ergs/cm2 per set) there is probably no independent effect on the b values of unacclimated and
884
ARTHUR L. BUIKEMA, JR.
acclimated animals nor is there a significant difference between adjusted means of unacclimated animals. However, acclimation to 1.7 ft-c did result in a significantly lower adjusted mean than that obtained for red wavelengths suggesting that a drop in average metabolic rate may be due to acclimation to a broad spectral range rather than to intensity or red wavelengths per se. DISCUSSION
Although the metabolism of crustaceans as a function of various environmental parameters has been studied frequently (Richman, 1958; Wolvekamp & Waterman, 1960) relatively few studies were concerned with the effect of these parameters on the relationship between rate of oxygen consumption and body size. Some workers have expressed their data as a function of body length (Obreshkove, 1930; Jancarik, 1948) while others have expressed their data relative to the metabolic rate of the first instar (Terao, 1931; Hoshi, 1949). The latter studies are not easily compared with other studies. The relationship between rate of oxygen consumption and body weight is described by the equation vol. 0, = aI&%,
(4)
where W is dry weight, b is the regression coefficient and a is the intercept value. The b value for crustacea generally varies between 0.67 and 1.00 (Wolvekamp & Waterman, 1960). A b value of 0.67 is interpreted to denote a relationship between metabolism and body surface area while a value of 1.00 has been described as a relationship between metabolism and body weight. Intermediate values, between 0.67 and l-00, are found for the majority of crustaceans. Weymouth et al. (1944) reported a mean b value of 0826 for a number of species of crustaceans and Zeuthen (1953) reported that the b value for organisms less than 1 mg nitrogen is O-95. Richman (1958) reported a b value of 0.881 for D. pulex. The b values obtained in this study are in general agreement when one compares the average b value determined for unacclimated animals, O-930 (0.979 if the data for 3.5 ft-c are included), and acclimated animals, O-780, although there is considerable variation among b values (Table 1). Interestingly, the b values are generally higher for unacclimated animals. Variation among b values is not unusual and they vary with temperature for snails (Berg & Ockelman, 1959) and amphipods (Armitage, 1962) and among various genera and/or species of marine copepods (Conover, 1959), crabs (Teal, 1959) and other crustaceans (Wolvekamp & Waterman, 1960). The results of this study suggest that light also influences the relationship between metabolism and body weight and that this relationship is also affected by light acclimation. The effect of light acclimation is more evident if the data obtained for the animals in the dark are compared to Richman’s data for the same species since Richman (1958) also tested his animals in the dark. The b value obtained by Richman (calculated from his data, 1958) is sigmficantly different from the b value obtained for unacclimated animals (F = 22.98, PC 0*005). Because the animals
OXYGEN
CONSUMPTION
OF
DAPHNIA
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885
used in this study were either not acclimated or acclimated for at least two complete molt cycles, the significant differences between my data and Richman’s are difficult to interpret. The significant difference in the b values may be an artifact, or it may be due to the fact that Richman’s animals were partially acclimated to the dark. Richman states that the animals were placed in filtered water 24 hr prior to the experiment but he does not state whether they were kept in the dark. He did adapt his animals to the dark in his feeding study. Because the adjusted means between animals not acclimated and acclimated to the dark are significantly different (this study) and also the adjusted mean between Richman’s data and that for acclimated animals is significantly different, it is possible that acclimation to the dark may be a type of translation. However, the significant difference between Richman’s data and mine for dark acclimated animals may be due to the fact that Richman’s animals were more crowded (fifty Da@& per 5 ml and 50-100/135 ml vs. five to twenty-five Duphniu per 60 ml) when he determined oxygen consumption. Crowding resulted in a lower rate of oxygen consumption for 5’. vet&s (Hoshi, 1957) but had an opposite effect on D. magna (Zeiss, 1963). If crowding caused an increase in oxygen consumption of D. pulex, I would have expected Richman’s results to be the same or higher than mine and they were not. If the data obtained in this experiment are expressed as ~1. O,/mg per hr, they compare well with those obtained by other authors as summarized by Richman (1958). As Richman noted, the weight specific rate of oxygen consumption of D. magna, Chydorus ovalis and C. Jinmarchicus are not in agreement and may be due to their larger size. Because light intensity is known to affect the activity of aquatic organisms (Yerkes, 1900; Dice, 1914; Schulz, 1928; Clarke, 1932; Ringelberg, 1963, 1964) one would expect that a change in metabolism should be demonstrable. Interestingly, Marshall et al. (1935) reported that the activity of C. jinmarchicus did not change when it was exposed to light even though the rate of oxygen consumption increased over 100 per cent. Activity was not observed in this experiment; but, because the swimming velocity of Duphniu increases as light intensity decreases, one might expect that metabolic rate would be higher at low light intensities. This was not necessarily the case (Fig. 1). Interestingly, for unacclimated animals there is no significant effect of light intensities above 14 ft-c on metabolic rate (Fig. 1). Marshall et al. (1935) reported an increased metabolic rate for C. Jinmarchicus when light intensity increased and that under subsequent increases in light intensity there was no effect, In acclimated organisms there is an increase in the rate of oxygen consumption above 14 ft-c. While the light intensities studied in this experiment are not very high, there is an effect of light increasing the metabolic rate of organisms, but the interpretation of the results depends on the light intensities being compared, acclimation state and size of the animals. In those organisms where there was no significant effect of light or dark on metabolic rate (Conover, 1956; Topping, 1965 ; Pearcy et al., 1969) the light intensities which were used may have
886
ARTHUR
L.
BUIKEMA,
JR.
coincidentally resulted in similar data and had other light intensities been used, the results may have been different. For example, the rates of oxygen consumption obtained for unacclimated, large animals (0.056 mg) in the dark, 28,55 and 110 ft-c are probably not significantly different even though the rate obtained for 14 ft-c is significantly lower (Fig. 1). Baylor & Smith (1953, 1957) suggest that the response of Daphnia to polarized light is due to an increase in the intensity of light which enters the cone lenses of the eye. The effect of polarized light (66 ft-c) on the b value was inseparable from the affect of 7 ft-c, but the adjusted means obtained for animals tested under polarized light are significantly different from that obtained at 7 ft-c but not from 14 ft-c (unacclimated) or 28 and 55 ft-c (acclimated) which suggests that the effect of polarized light on the rate of oxygen consumption may be due to an apparent higher intensity perceived by the animal. Previously cited studies suggest that wavelengths, because they affect activity, may also affect the rate of oxygen consumption. The results obtained in this experiment do not support such a hypothesis (Fig. 2). Even though there is a considerable difference in the nature and velocity of the blue dance, which is associated with food-hunting, and the red dance, which maintained the organism in an area of high food concentration (Baylor & Smith, 1957; Smith & Baylor, 1959) it is suggested that a Daphnia which is maintaining its position in an area of high food concentration expends as much energy as a Daphnia searching for food. SUMMARY 1.
The effects of nine light intensities (0, 1.7, 3.5, 7, 14, 28, 55 and 110 ft-c), polarized light (6.6 ft-c) and four wavelength ranges were studied on the relationship (b) of metabolic rate and body size for unacclimated and acclimated Daphnia pulex. 2. Light intensity significantly affects b and the effect varies with acclimation. Light intensities of 1.7 and 3.5 ft-c were stimulatory, 14 ft-c was depressant and intensities above 28 ft-c were slightly depressant on the metabolic rate of different sized unacclimated animals. Light intensities of 1.7 and 28 ft-c were stimulatory and intensities of 3.5, 7 and 14 ft-c were depressant on the metabolic rate of young animals and the effects were reversed in older animals. Above 14 ft-c the metabolic rate of older acclimated animals increases linearly with light intensity. 3. The effect of polarized light on b was inseparable from the effect of light intensity but there were significant differences among adjusted means. Acclimation to polarized light resulted in a significantly higher adjusted mean. 4. There was no significant effect of wavelength among b values, and adjusted mean values, for unacclimated and acclimated animals. Acclimation to each wavelength did result in a significantly lower b value. Acknowledgements-I encouragement
am indebted to my major and co-operation during this study.
advisor,
Dr.
K.
B. Armitage,
for his
OXYGEN
CONSUMPTION
OF DAPHNIA
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consumption ; cladocera ; Daphnia pulex ; light ; acclimation ;