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Water balance in Collembola and its relation to habitat selection: Water content, haemolymph osmotic pressure and transpiration during an instar
J. Insect Physiol., Vol. 21, No. 11, pp. 755-760, 1981 Printed in Great Britain.
0022-1910/81/l 107554I6$02.00/0 0 1981 Pergumon Press Ltd.
WATER BALANCE IN COLLEMBOLA AND ITS RELATION TO HABITAT SELECTION: WATER CONTENT, HAEMOLYMPH OSMOTIC PRESSURE AND TRANSPIRATION DURING AN INSTAR H. A. VERHOEF Department of Biology, Vrije Universiteit, De Boelelaan 1087, Amsterdam-Buitenveldert. The Netherlands (Received 20 February 1981; revised 24 April 1981) AMrae-In the two collembolan species Orchesellu cincta and Tomocerus minor the water content, haemolymph osmotic pressure and transpiration rate fluctuate with the feeding rhythm during each instar. The changes in water content, however, are due to changes in dry weight, because the absolute water weight stays constant during the instar. The intake of food is probably the cause of the increase in haemolymph osmotic pressure. Increase of osmotically active substances in the blood and/or blood volume reduction may be responsible for the rise in osmotic pressure. This change in osmotic pressure in turn may affect the responsiveness of the animals to water as well as their feeding behaviour. Changes in the epicuticle and in epidermal cell membranes may cause changes in the rate of transpiration. The high rate observed during ecdysis and during the mid-instar may explain the behaviour of the animals in varied water conditions. Dehydration during the instar causes an equivalent rise in osmotic pressure for both Tomocerus minor and Orchesellu cinctu. The water loss appears to involve the haemolymph. The physiological state of the animal does not influence the rise in osmotic pressure. There are no signs of any osmoregulation in the two species. Key Word Index: Water balance, Collembola, haemolymph osmotic pressure, transpiration rate, habitat
INTRODUCTION IN A previous study on the water relations in the coexisting species of Collembola Orchesella cincta (L.) and Tomocerus minor (Lubbock), important differences are found in the integumentary water loss and water uptake of these animals. The respective transpiration rates are 143 and 581 pg cmm2 hr- l mm Hg - *, whereas the water uptake rates are 12 and 2 pg min-1 (VERHOEF and WITTEVEEN, 1980). These differences in output and input of water partly explain the different reactions of these two species to moisture conditions and the different micro-distribution in their natural habitat: T. minor inhabits the moist parts of the habitat, whereas 0. cincta can also be found in the drier areas. However, the reaction of the species to moisture conditions changes throughout life and seems to be influenced by the various life stages (VERHOEF and NAGELKERKE, 1977). The life cycle of these frequently-moulting animals is divided into many instars, each of which is divided into a feeding period and a period in which feeding is impossible due to degeneration of parts of the gut-epithelium (VERHOEF, unpublished observations). During about 60”,/, of the total duration of the instar (4-5 days at 2O”C), these species have a full gut (JOOSSE and TFSTERINK, 1977). In adults, reproduction takes place during alternate instars immediately after ecdysis. An experimental study on the spatial distribution of these animals in a heterogenous water/food situation madeitclearthat thevariouslifestagesgreatlyinfluence
the distribution of the animals: during ecdysis and subsequent reproduction 0. cincta assembles in watersaturated places. This leads to dense contact aggregation, probably caused by short-distance active aggregation pheromones (VERHOEF et al., 1977). ‘Dispersal’ follows during the feeding period. After feeding, orientation towards water-saturated places occurs by means of orthokinetic reactions and the aggregations are re-established. This ‘periodical aggregation’ on wet places is not found for T. minor. Under the moisture conditions given, this species appears to be totally restricted in distribution to waterwhere it forms spaced-out saturated places, aggregations (VERHOEF and NAGELKERKE, 1977). To clarify whether changes in water conditions of these animals during the instar may be responsible for the different reactions of the animals to the water situation of their habitat, three important parameters have been estimated during the instar: body water content, blood osmotic pressure of hydrated and dehydrated animals, and transpiration. The results are reported in this paper.
MATERIALS
AND METHODS
The springtail species used in this study are the epedaphic species, Orchesella cincta and Tomocerus minor, both collected from a mixed birch-pine forest near Hilversum, the Netherlands. For a description of the methods of transpiration rate and blood osmotic 155
l
H. A. VERHOEF
156
pressure measurements, see VERHOEFand WITTEVEEN (1980). In order to establish the moulting stage and the feeding stage, each individual was observed 3 times a day. The animals were kept in separate boxes (diameter 4.5 cm) with a base of plaster of Paris (which was kept damp by distilled water) in a climate room at 20°C. As food, a suspension of Pleurococcus sp. (green algae) was offered. For the water content measurements, 20 animals were used for each species. Fresh weight was determined daily and 4 individuals were separated for dry weight measurement. Thus, at the end of the fiveday instar. 4 animals were left for fresh and subsequently dry weight measurements. Dry weight was obtained by drying over silica gel to a constant weight at 60°C. The water content was expressed as a percentage of wet weight. For the osmotic pressure measurements in 0. cincra, haemolymph samples were taken both at the antennae (see VERHOEF and WITTEVEEN,1980) and at the ventral vesicles, to see if there are any local differences in the osmotic pressure of the haemolymph of this species. After haemolymph sampling in both species the animal was cut open ventrally to study the gut contents in order to get a more exact indication of the feeding period. Dehydration in both species was performed at 38”” r.h. and 20°C. The effect of a 10% weight loss on the blood osmotic pressure was measured for both species. Due to their different transpiration rates (see VERHOEF and WITTEVEEN,1980) the desiccation time for T. minor was 20 min and for 0. cincta 60 min. The expected osmotic pressure after dehydration was calculated on the assumption that, in the absence of osmoregulation, water loss causes an equivalent rise in osmotic pressure (cf. WOODRING,1974). For transpiration measurements, 5 individuals were used per species. Transpiration was measured daily at Weight
fmgl
Tomocerus
I
minor
10.00 a.m. at 0;; r.h. and 20°C. Afterwards the animals were replaced in the boxes with ample water and food. Twenty-four hours later all animals were fully rehydrated (VERHOEF and WITTEVEEN,1980). The water loss was expressed as a percentage of the fresh weight of the individual animal concerned. As it is impossible to estimate the gut content of the living individual during the instar, changes in fresh weight were used to indicate the feeding period: the end of the weight increase was taken as the end of the feeding period.
RESULTS Water content
The water content of both 0. cinctu and T. minor expressed as a percentage of fresh weight showed a decrease during the first 4 days of the instar followed by an increase on the last day (see Fig. 1). However, both fresh and dry weight increased and decreased equally; thus the absolute weight of water remained more or less constant during an instar. Concerning fresh weight Fig. 1 shows for 0. cincta the main increase in the first 2 days; for T. minor there is a more continuous increase. Haemolymph osmotic pressure
The course in osmotic pressure of the haemolymph during the instar, given in Fig. 2, showed a maximum of about 350 mosm for 0. cincta on the third day of the instar. In T. minor the highest value of 325 mosm was found on both the second and the third day. In 0. cincta the osmotic pressure of the haemolymph taken from the ventral vesicles showed no difference from the values of the haemolymph taken from the antennae, so there are probably no local differences in the osmotic pressure of the blood of this species.
I
Orchesello cincta
0.400
0.200
*&ii+>, I
141
Water content 80 (% fresh we,ght J !;\
70
‘:h$ f4J
i r
-*
I
1 Fig. 1. Fresh (+o).
2
,
3
(3,
I
4
I
5
*
Y-• 7
2
3
dry (A-A) and water weight (o-o) and water content minor and 0. cincra during an instar (+ S.E.M.) * = ecdysis.
4 (U---m)
5
+days
of 7’
Water
balance
in Collembola
and its relation
Hoemolymph 350 - osmotic pressure fmosmol
//ter
to habitat
selection
151
Table 1. Analysis of variance of the rise in osmotic pressure due to dehydration in Orchesella cincia and Tomocerus minor. The analysis evaluates the effects of physiological state and species
-I!
340 Source
Physiological Species Error Total
320-
300
of variation
-
DF
MS
F
4 1 4
151.23 638,40 94.85 1622.73
1.59 ns. 6.73 ns.
state
n.s.: not significant at 57: probability level. DF = degrees of freedom; MS = mean square: variance ratio.
F =
cincta
26Ot
-s
1
2
Fig. 2. Haemolymph osmolarity during an instar (If: S.E.M.).
3
4
5
+-
days
of 0. cincta and T. minor
Number in parentheses = number of test animals; * = ecdysis.
the same occurred 30 min after ecdysis. Concerning the course of transpiration during the instar, it can be concluded that transpiration varies in relation to the feeding rhythm within a relatively small range. Effect
Transpiration
The results for transpiration are given in Fig. 3. There are common differences in time between the graphs for the individual animals since they were not exactly at the same moulting stage. Both species showed a relatively high transpiration rate at the end of the feeding period. Maximal values were reached immediately (10 set) after ecdysis. Sixty minutes after exdysis, the transpiration of the fully rehydrated animals was back at its ‘pre-ecdysis’ level. For 0.
of dehydration on haemolymph
osmotic pressure
The effects of a 10% weight loss on the haemolymph osmotic pressure are given in Fig. 4. In both T. minor and 0. cincta this weight loss caused a significant rise (PI 0.01, Mann-Whitney U-test). The expected osmotic pressure of dehydrated animals is also given in Fig. 4. For both species these values lay within the 95% confidence limits of the means of the observed data. Thus the water loss is primarily from the haemolymph. The effect of the physiological state (:day of the instar) on the rise in osmotic pressure due to Orchesello Tronspwatmn (% body weight loss/ lOminI
oncta
5.00 -
4.w-
300-
soo-
4.00 -
3.00 -
2.00 -
aA COO-
4.00-
300-
8.00 -
iL
200.
,&JL--I)
1
1
8
2
8
3
*
4
+A
5
,OOL--*
days
I
1
3
2
i.
-3
l2
a
6
3
4
5
days
Fig. 3. Transpiration of 0. cincta and T. minor during an instar, shown for 3 specimens per species. The arrow indicates the end of the feeding period. * = ecdysis; a = 10 set after ecdysis; b = 30 min after ecdysis; c = 60 min after ecdysis.
H. A. VERHOEF
158 Tomocerus Hoemolymph pressure
minor
Urchesello
cincto
osmotic
lmosmolliter-l)
340-
320-
300
-
280-
260
t
--+
,
’
l
2
3
4
5
-
-*
1
2
3
4
5
*-
days
Fig. 4. Haemolymph osmolarity df T. minor.fully hydrated (A-A) and dehydrated (a-a) and 0. &cm, fully hydrated (t_@)and dehydrated (+o), and the expected osmolarity after dehydration (0 0) during an instar ( f S.E.M.). Number in parentheses = number of test animals; * = ecdysis.
dehydration in 0. cincta and T. minor is given in Table 1. It appeared that the physiological state of the animal did not influence the rise in osmotic pressure. In this respect there was no difference between the two species. DISCUSSION The water content of Orchesella cincta and that of Tomocerus minor, expressed as average percentages of the total body weight, were about the same: 72.5-73.5x. As 0. cincta is considered to be the more terrestrially adopted species of the two (VERHOEFand WITTEVEEN, 1980), the opinion of HOLDICH and MAYES (1976). that a relatively low water content is part of an adaptation to terrestrial life does not hold for these two species. During the instar, the water content of both species fluctuated. The absolute weight, however, stayed constant during the instar (Fig. 1). This was found also in T. minor after a short starvation period of three days in which the decrease in fresh weight equalled the decrease in dry weight and the water content remained constant (VANNIER and VERHOEF, 1978). Both effects were found in the thysanuran Thermobia domestica Packard by OKASHA (1971), who refers to “volume regulation”. The average osmotic pressure of the haemolymph was for 0. cincta 320 and for T. minor 310 mosm litre-l, values fitting in the range of 300-500 mosm litre - ’ given for insects (EDNEY, 1977). In both collembolan species the blood osmotic pressure appeared to fluctuate during the instar: it increased in
the feeding period with 30 (T. minor) and 45 mosm (0. cincta)anddecreasedinthesecondhalfoftheinstarwith 41 and 73 mosm respectively. This might be associated with feeding as has been reported in Locusta (BERNAYS and CHAPMAN, 1974). In this case the effect of feeding was a short-period reduction of the blood volume, which was reflected in a rise of about 50 mosm in blood osmotic pressure. Furthermore, mid-instar drops in blood volume have been found in the locust the hemipteran Schistocerca (LEE, 1961) and Dysdercus (BERRIDGE, 1965). Fluctuations in the blood osmotic pressure of locusts have been shown to affect the responsiveness of the animals to water and their feeding behaviour (BERNAYSand CHAPMAN, 1974; BARTON BROWNE and VAN GERWEN, 1976). The observed mid-instar rise of blood osmotic pressure in 0. cincta might be a stimulus to search for wet places after feeding (see VERHOEFand NAGELKERKE, 1977). Rise of blood osmotic pressure is also caused by dehydration. The haemolymph can be considered to be the major water reservoir. Any form of osmoregulation seems to be absent both in the drought-tolerant 0. cincta and in the more droughtsensitive species T. minor. In the field, this kind of rapid dehydration may occur in heterogeneous moisture conditions when the animals reach dry areas by a random walk. Both species appear to react by a behavioural mechanism, i.e. an orthokinetic reaction by which they reach the wet areas again (VERHOEFand NAGELKERKE, 1977). During slow dehydration, in dry summers when the litter dries out slowly, 0. cincta seems to be able to regulate its water content and
Water
balance
in Collembola and its relation to habitat selection
blood osmotic pressure to some extent (VERHOEF, unpublished). The average transpiration rate for 0. cincta is about 2.5% and for T. minor about 8.8% body weight loss 10 min-I. This large difference in average water loss explains the difference in water dependence between the two species. During the instar a fluctuation similar to that found for the blood osmotic pressure appeared to exist in the transpiration of both species: the mid-instar rise in transpiration coincided with the end of the feeding period. These changes might be caused by changes in the permeability of the barriers against integumentary water loss: the epicuticle and/or the epidermal cell membranes (BERRIDGE, 1970; TREHERNE and WILLMER, 1975). Changes in the epicuticle can be caused by abrasion as was found for the collembolan Podura aquatica (NOBLE-NESBITT, 1963); the presence and composition of the epicuticular lipids may also change as found for ticks (DAVIS, 1974) and the tenebrionid Eleodes armata (HADLEY, 1977). The importance of the epicuticular lipids is shown clearly in the increased transpiration during ecdysis, as hasbeen found in several arthropods, e.g. the collembolan Seira domestica (VANNIER. 1973). It has been assumed that the waterproofing lipid-layer does not reach its full impermeability for some time (BEAMENT, 1976; BR~~cK, 1976). The presence of the animals on water-saturated areas during ecdysis may be related to the permeable state of their cuticle. Furthermore, in two respects free water is essential for a successful moult: a high water content is necessary to generate the hydrostatic pressure for the splitting of the exuvia, whereas newly moulted animals need water for initial growth since at that time the cuticle is visibly soft and flexible. The same applies to the eggs. The cuticle of new-laid eggs is very permeable and the uptake of water for the initial growth is important for successful development (HALE, 1965). The mid-instar rise in transpiration might induce an orthokinetic locomotory reaction causing orientation towards saturated places (JOOSSEand VERHOEF, 1974). It might also be possible that the high transpiration forms a way for the elimination of water at the end of the feeding period, as the two species concerned fed on a wet Pleurococcus diet. According to SUTTON (1972) the more drought-tolerant isopods appear to transpire the excess of water. This method is more efficient than an equivalent output by the excretory system, which would necessarily be associated with a loss of physiologically valuable solutes and would involve a considerable expenditure of energy in ion transport. This mid-instar rise in transpiration rate is probably no active enhancement of the transpiration rate, as the same rise occurs in 0. cirzcta and T. minor alike. The latter species seems to be unable to actively influence the transpiration rate (VERHOEF and WITTEVEEN. 1980). Ach-nowlerlgements-The author is grateful to Drs. E. N. G. J~~SSE and N. D. DE WITH for helpful discussions, to Dr. J. M. SKELDING, Westfield College, University of London, for the introduction into the technique of measuring haemolymph osmolarity, to Mr. G. W. H. VAN DEN BERG for drawing the figures and to Miss. S. RICHTER for typing the manuscript.
759
REFERENCES BARTON BROWNE L. and VAN GERWEN A. C. M. (1976) Regulation of water ingestion by the locust, Chortoicetes terminifera: the effect of injections into the haemolymph. Phvsiol. Em. 1, 159-167. BEAMENTJ. W. L. (1976) The ecology of cuticle. In The Insect Integument (Ed. by HEPBURN H. R. ), pp. 359-374. Elsevier, Oxford. BERNAYS E. A. and CHAPMAN R. F. (1974) The effect of haemolymph osmotic pressure on the meal size of nymphs of Locusta migratoria L. J. exp. Biol. 61, 473480. BERRIDGE M. J. (1965) The physiology of excretion in the cotton stainer, Dysdercus fasciatus Signoret. II. Inorganic excretion and ionic regulation. J. exp. Biol. 43; 523-533. BERRIDGE M. J. (1970) Osmoregulation in terrestrial arthropods. In Chemical Zoology (Ed. by FLORKIN M. and SCHEERB. T.) Vol. V. part A, pp. 287-320. Academic Press, New York. BR~~CK E. (1976) Die Durchlsssigkeit der Cuticula von Blaberus trapezoideus und Periplaneta americana fiir eine wlssrige anorganische PhosphatlBsung (Blattaria: Blaberidae/Blattidae). Ent. Germ. 2, 320-327. DAVIS M. T. B. (1974) Critical temperature and changes in cuticular lipids in the rabbit tick, Haemaphysalis leporispalustris. J. Insect Physiol. 20, 1087-l 100. EDNEY E. B. (1977) Water Balance in Land Arthropods. Springer, Berlin. HADLEY N. F. (1977) Epicuticular lipids of the desert tenebrionid beetle, Eleodes armata: seasonal and acclimatory effects on composition. Insect Biochem. 7, 277-283. HALE W. G. (1965) Observations on the breeding biology of Collembola. Pedobiologia 5, 161-177. HOLDICH D. M. and MAYE_SK. R. (1976) Blood volume and total water content of the woodlouse, Oniscus asellus, in conditions of hydration and desiccation. J. Insect Ph.vsiol. 22, 547-553. JOOSSEE. N. G. and TFXERINK G. J. (1977) The role of food in the population dynamics of Orchesella cincta (Linnt) (Collembola). Oecologia (Berl.) 29, 189-204. J~IXSE E. N. G. and VERHOEF H. A. (1974) On the aggregational habits of surface dwelling Collembola. Pedobiologia 14, 245-249. LEE R. M. (1961) The variation of blood volume with age in the desert locust (Schistocerca gregaria Forsk.). J. Insect Physiol. 6, 36-5 1. NOBLE-NESBITTJ. (1963) Transpiration in Podura aquatica L. (Collembola, Isotomidae) and the wetting properties of its cuticle. J. exp. Biol. 40, 681-700. OKASHA A. Y. K. (1971) Water relations in an insect, Thermobia domestica. I. Water uptake from sub-saturated atmospheres as a means of volume regulation. J. e.xp. Biol. 55,435448. SUTTON S. L. (1972) Woodlice. Ginn, London. TREHERNEJ. E. and WILLMER P. G. (1975) Hormonal control of integumentary water-loss: evidence for a novel neuroendocrine system in an insect (Periplaneta americana). J. exp. Biol. 63, 143-159. VANNIER G. (1973) Etude de la transpiration chez un Insecte Collembole au tours de son exuviation. C.r. hebd. Skanc. Acad. Sci. Paris 211, 2231-2234. VANNIER G. and VERHOEFH. A. (1978). Effect of starvation on transpiration and water content in the populations of two coexisting Collembola species. Camp. Biochem. Physiol. 6OA, 483489. VERHOEF H. A. and WIT~EVEENJ. (1980) Water balance in Collembola and its relation to habitat selection; cuticular water loss and water uptake. J. Insect Physiol. 26,201-208. VERHOEFH. A. and NAGELKERKEC. J. (1977) Formation and ecological significance of aggregations in Collembola; an experimental study. Oecologia (Bed.) 31, 215-226. VERHOEFH. A., NAGELKERKEC. J. and JOOSSE E. N. G. (1977)
760
H. A. VERHOEF
Aggregation pheromones in Collembola. J. Insecf Physiol. 23, 1009-1013. WOODRINGJ. P. (1974) Effects of rapid and slow dehydration
on the haemolymph osmolarity and Na+-K+ concentration in the millipede Pachydesmus crassicutis. Camp. Biochem. Physiol. 49A, 115-I 19.
0022-1910/81/l 107554I6$02.00/0 0 1981 Pergumon Press Ltd.
WATER BALANCE IN COLLEMBOLA AND ITS RELATION TO HABITAT SELECTION: WATER CONTENT, HAEMOLYMPH OSMOTIC PRESSURE AND TRANSPIRATION DURING AN INSTAR H. A. VERHOEF Department of Biology, Vrije Universiteit, De Boelelaan 1087, Amsterdam-Buitenveldert. The Netherlands (Received 20 February 1981; revised 24 April 1981) AMrae-In the two collembolan species Orchesellu cincta and Tomocerus minor the water content, haemolymph osmotic pressure and transpiration rate fluctuate with the feeding rhythm during each instar. The changes in water content, however, are due to changes in dry weight, because the absolute water weight stays constant during the instar. The intake of food is probably the cause of the increase in haemolymph osmotic pressure. Increase of osmotically active substances in the blood and/or blood volume reduction may be responsible for the rise in osmotic pressure. This change in osmotic pressure in turn may affect the responsiveness of the animals to water as well as their feeding behaviour. Changes in the epicuticle and in epidermal cell membranes may cause changes in the rate of transpiration. The high rate observed during ecdysis and during the mid-instar may explain the behaviour of the animals in varied water conditions. Dehydration during the instar causes an equivalent rise in osmotic pressure for both Tomocerus minor and Orchesellu cinctu. The water loss appears to involve the haemolymph. The physiological state of the animal does not influence the rise in osmotic pressure. There are no signs of any osmoregulation in the two species. Key Word Index: Water balance, Collembola, haemolymph osmotic pressure, transpiration rate, habitat
INTRODUCTION IN A previous study on the water relations in the coexisting species of Collembola Orchesella cincta (L.) and Tomocerus minor (Lubbock), important differences are found in the integumentary water loss and water uptake of these animals. The respective transpiration rates are 143 and 581 pg cmm2 hr- l mm Hg - *, whereas the water uptake rates are 12 and 2 pg min-1 (VERHOEF and WITTEVEEN, 1980). These differences in output and input of water partly explain the different reactions of these two species to moisture conditions and the different micro-distribution in their natural habitat: T. minor inhabits the moist parts of the habitat, whereas 0. cincta can also be found in the drier areas. However, the reaction of the species to moisture conditions changes throughout life and seems to be influenced by the various life stages (VERHOEF and NAGELKERKE, 1977). The life cycle of these frequently-moulting animals is divided into many instars, each of which is divided into a feeding period and a period in which feeding is impossible due to degeneration of parts of the gut-epithelium (VERHOEF, unpublished observations). During about 60”,/, of the total duration of the instar (4-5 days at 2O”C), these species have a full gut (JOOSSE and TFSTERINK, 1977). In adults, reproduction takes place during alternate instars immediately after ecdysis. An experimental study on the spatial distribution of these animals in a heterogenous water/food situation madeitclearthat thevariouslifestagesgreatlyinfluence
the distribution of the animals: during ecdysis and subsequent reproduction 0. cincta assembles in watersaturated places. This leads to dense contact aggregation, probably caused by short-distance active aggregation pheromones (VERHOEF et al., 1977). ‘Dispersal’ follows during the feeding period. After feeding, orientation towards water-saturated places occurs by means of orthokinetic reactions and the aggregations are re-established. This ‘periodical aggregation’ on wet places is not found for T. minor. Under the moisture conditions given, this species appears to be totally restricted in distribution to waterwhere it forms spaced-out saturated places, aggregations (VERHOEF and NAGELKERKE, 1977). To clarify whether changes in water conditions of these animals during the instar may be responsible for the different reactions of the animals to the water situation of their habitat, three important parameters have been estimated during the instar: body water content, blood osmotic pressure of hydrated and dehydrated animals, and transpiration. The results are reported in this paper.
MATERIALS
AND METHODS
The springtail species used in this study are the epedaphic species, Orchesella cincta and Tomocerus minor, both collected from a mixed birch-pine forest near Hilversum, the Netherlands. For a description of the methods of transpiration rate and blood osmotic 155
l
H. A. VERHOEF
156
pressure measurements, see VERHOEFand WITTEVEEN (1980). In order to establish the moulting stage and the feeding stage, each individual was observed 3 times a day. The animals were kept in separate boxes (diameter 4.5 cm) with a base of plaster of Paris (which was kept damp by distilled water) in a climate room at 20°C. As food, a suspension of Pleurococcus sp. (green algae) was offered. For the water content measurements, 20 animals were used for each species. Fresh weight was determined daily and 4 individuals were separated for dry weight measurement. Thus, at the end of the fiveday instar. 4 animals were left for fresh and subsequently dry weight measurements. Dry weight was obtained by drying over silica gel to a constant weight at 60°C. The water content was expressed as a percentage of wet weight. For the osmotic pressure measurements in 0. cincra, haemolymph samples were taken both at the antennae (see VERHOEF and WITTEVEEN,1980) and at the ventral vesicles, to see if there are any local differences in the osmotic pressure of the haemolymph of this species. After haemolymph sampling in both species the animal was cut open ventrally to study the gut contents in order to get a more exact indication of the feeding period. Dehydration in both species was performed at 38”” r.h. and 20°C. The effect of a 10% weight loss on the blood osmotic pressure was measured for both species. Due to their different transpiration rates (see VERHOEF and WITTEVEEN,1980) the desiccation time for T. minor was 20 min and for 0. cincta 60 min. The expected osmotic pressure after dehydration was calculated on the assumption that, in the absence of osmoregulation, water loss causes an equivalent rise in osmotic pressure (cf. WOODRING,1974). For transpiration measurements, 5 individuals were used per species. Transpiration was measured daily at Weight
fmgl
Tomocerus
I
minor
10.00 a.m. at 0;; r.h. and 20°C. Afterwards the animals were replaced in the boxes with ample water and food. Twenty-four hours later all animals were fully rehydrated (VERHOEF and WITTEVEEN,1980). The water loss was expressed as a percentage of the fresh weight of the individual animal concerned. As it is impossible to estimate the gut content of the living individual during the instar, changes in fresh weight were used to indicate the feeding period: the end of the weight increase was taken as the end of the feeding period.
RESULTS Water content
The water content of both 0. cinctu and T. minor expressed as a percentage of fresh weight showed a decrease during the first 4 days of the instar followed by an increase on the last day (see Fig. 1). However, both fresh and dry weight increased and decreased equally; thus the absolute weight of water remained more or less constant during an instar. Concerning fresh weight Fig. 1 shows for 0. cincta the main increase in the first 2 days; for T. minor there is a more continuous increase. Haemolymph osmotic pressure
The course in osmotic pressure of the haemolymph during the instar, given in Fig. 2, showed a maximum of about 350 mosm for 0. cincta on the third day of the instar. In T. minor the highest value of 325 mosm was found on both the second and the third day. In 0. cincta the osmotic pressure of the haemolymph taken from the ventral vesicles showed no difference from the values of the haemolymph taken from the antennae, so there are probably no local differences in the osmotic pressure of the blood of this species.
I
Orchesello cincta
0.400
0.200
*&ii+>, I
141
Water content 80 (% fresh we,ght J !;\
70
‘:h$ f4J
i r
-*
I
1 Fig. 1. Fresh (+o).
2
,
3
(3,
I
4
I
5
*
Y-• 7
2
3
dry (A-A) and water weight (o-o) and water content minor and 0. cincra during an instar (+ S.E.M.) * = ecdysis.
4 (U---m)
5
+days
of 7’
Water
balance
in Collembola
and its relation
Hoemolymph 350 - osmotic pressure fmosmol
//ter
to habitat
selection
151
Table 1. Analysis of variance of the rise in osmotic pressure due to dehydration in Orchesella cincia and Tomocerus minor. The analysis evaluates the effects of physiological state and species
-I!
340 Source
Physiological Species Error Total
320-
300
of variation
-
DF
MS
F
4 1 4
151.23 638,40 94.85 1622.73
1.59 ns. 6.73 ns.
state
n.s.: not significant at 57: probability level. DF = degrees of freedom; MS = mean square: variance ratio.
F =
cincta
26Ot
-s
1
2
Fig. 2. Haemolymph osmolarity during an instar (If: S.E.M.).
3
4
5
+-
days
of 0. cincta and T. minor
Number in parentheses = number of test animals; * = ecdysis.
the same occurred 30 min after ecdysis. Concerning the course of transpiration during the instar, it can be concluded that transpiration varies in relation to the feeding rhythm within a relatively small range. Effect
Transpiration
The results for transpiration are given in Fig. 3. There are common differences in time between the graphs for the individual animals since they were not exactly at the same moulting stage. Both species showed a relatively high transpiration rate at the end of the feeding period. Maximal values were reached immediately (10 set) after ecdysis. Sixty minutes after exdysis, the transpiration of the fully rehydrated animals was back at its ‘pre-ecdysis’ level. For 0.
of dehydration on haemolymph
osmotic pressure
The effects of a 10% weight loss on the haemolymph osmotic pressure are given in Fig. 4. In both T. minor and 0. cincta this weight loss caused a significant rise (PI 0.01, Mann-Whitney U-test). The expected osmotic pressure of dehydrated animals is also given in Fig. 4. For both species these values lay within the 95% confidence limits of the means of the observed data. Thus the water loss is primarily from the haemolymph. The effect of the physiological state (:day of the instar) on the rise in osmotic pressure due to Orchesello Tronspwatmn (% body weight loss/ lOminI
oncta
5.00 -
4.w-
300-
soo-
4.00 -
3.00 -
2.00 -
aA COO-
4.00-
300-
8.00 -
iL
200.
,&JL--I)
1
1
8
2
8
3
*
4
+A
5
,OOL--*
days
I
1
3
2
i.
-3
l2
a
6
3
4
5
days
Fig. 3. Transpiration of 0. cincta and T. minor during an instar, shown for 3 specimens per species. The arrow indicates the end of the feeding period. * = ecdysis; a = 10 set after ecdysis; b = 30 min after ecdysis; c = 60 min after ecdysis.
H. A. VERHOEF
158 Tomocerus Hoemolymph pressure
minor
Urchesello
cincto
osmotic
lmosmolliter-l)
340-
320-
300
-
280-
260
t
--+
,
’
l
2
3
4
5
-
-*
1
2
3
4
5
*-
days
Fig. 4. Haemolymph osmolarity df T. minor.fully hydrated (A-A) and dehydrated (a-a) and 0. &cm, fully hydrated (t_@)and dehydrated (+o), and the expected osmolarity after dehydration (0 0) during an instar ( f S.E.M.). Number in parentheses = number of test animals; * = ecdysis.
dehydration in 0. cincta and T. minor is given in Table 1. It appeared that the physiological state of the animal did not influence the rise in osmotic pressure. In this respect there was no difference between the two species. DISCUSSION The water content of Orchesella cincta and that of Tomocerus minor, expressed as average percentages of the total body weight, were about the same: 72.5-73.5x. As 0. cincta is considered to be the more terrestrially adopted species of the two (VERHOEFand WITTEVEEN, 1980), the opinion of HOLDICH and MAYES (1976). that a relatively low water content is part of an adaptation to terrestrial life does not hold for these two species. During the instar, the water content of both species fluctuated. The absolute weight, however, stayed constant during the instar (Fig. 1). This was found also in T. minor after a short starvation period of three days in which the decrease in fresh weight equalled the decrease in dry weight and the water content remained constant (VANNIER and VERHOEF, 1978). Both effects were found in the thysanuran Thermobia domestica Packard by OKASHA (1971), who refers to “volume regulation”. The average osmotic pressure of the haemolymph was for 0. cincta 320 and for T. minor 310 mosm litre-l, values fitting in the range of 300-500 mosm litre - ’ given for insects (EDNEY, 1977). In both collembolan species the blood osmotic pressure appeared to fluctuate during the instar: it increased in
the feeding period with 30 (T. minor) and 45 mosm (0. cincta)anddecreasedinthesecondhalfoftheinstarwith 41 and 73 mosm respectively. This might be associated with feeding as has been reported in Locusta (BERNAYS and CHAPMAN, 1974). In this case the effect of feeding was a short-period reduction of the blood volume, which was reflected in a rise of about 50 mosm in blood osmotic pressure. Furthermore, mid-instar drops in blood volume have been found in the locust the hemipteran Schistocerca (LEE, 1961) and Dysdercus (BERRIDGE, 1965). Fluctuations in the blood osmotic pressure of locusts have been shown to affect the responsiveness of the animals to water and their feeding behaviour (BERNAYSand CHAPMAN, 1974; BARTON BROWNE and VAN GERWEN, 1976). The observed mid-instar rise of blood osmotic pressure in 0. cincta might be a stimulus to search for wet places after feeding (see VERHOEFand NAGELKERKE, 1977). Rise of blood osmotic pressure is also caused by dehydration. The haemolymph can be considered to be the major water reservoir. Any form of osmoregulation seems to be absent both in the drought-tolerant 0. cincta and in the more droughtsensitive species T. minor. In the field, this kind of rapid dehydration may occur in heterogeneous moisture conditions when the animals reach dry areas by a random walk. Both species appear to react by a behavioural mechanism, i.e. an orthokinetic reaction by which they reach the wet areas again (VERHOEFand NAGELKERKE, 1977). During slow dehydration, in dry summers when the litter dries out slowly, 0. cincta seems to be able to regulate its water content and
Water
balance
in Collembola and its relation to habitat selection
blood osmotic pressure to some extent (VERHOEF, unpublished). The average transpiration rate for 0. cincta is about 2.5% and for T. minor about 8.8% body weight loss 10 min-I. This large difference in average water loss explains the difference in water dependence between the two species. During the instar a fluctuation similar to that found for the blood osmotic pressure appeared to exist in the transpiration of both species: the mid-instar rise in transpiration coincided with the end of the feeding period. These changes might be caused by changes in the permeability of the barriers against integumentary water loss: the epicuticle and/or the epidermal cell membranes (BERRIDGE, 1970; TREHERNE and WILLMER, 1975). Changes in the epicuticle can be caused by abrasion as was found for the collembolan Podura aquatica (NOBLE-NESBITT, 1963); the presence and composition of the epicuticular lipids may also change as found for ticks (DAVIS, 1974) and the tenebrionid Eleodes armata (HADLEY, 1977). The importance of the epicuticular lipids is shown clearly in the increased transpiration during ecdysis, as hasbeen found in several arthropods, e.g. the collembolan Seira domestica (VANNIER. 1973). It has been assumed that the waterproofing lipid-layer does not reach its full impermeability for some time (BEAMENT, 1976; BR~~cK, 1976). The presence of the animals on water-saturated areas during ecdysis may be related to the permeable state of their cuticle. Furthermore, in two respects free water is essential for a successful moult: a high water content is necessary to generate the hydrostatic pressure for the splitting of the exuvia, whereas newly moulted animals need water for initial growth since at that time the cuticle is visibly soft and flexible. The same applies to the eggs. The cuticle of new-laid eggs is very permeable and the uptake of water for the initial growth is important for successful development (HALE, 1965). The mid-instar rise in transpiration might induce an orthokinetic locomotory reaction causing orientation towards saturated places (JOOSSEand VERHOEF, 1974). It might also be possible that the high transpiration forms a way for the elimination of water at the end of the feeding period, as the two species concerned fed on a wet Pleurococcus diet. According to SUTTON (1972) the more drought-tolerant isopods appear to transpire the excess of water. This method is more efficient than an equivalent output by the excretory system, which would necessarily be associated with a loss of physiologically valuable solutes and would involve a considerable expenditure of energy in ion transport. This mid-instar rise in transpiration rate is probably no active enhancement of the transpiration rate, as the same rise occurs in 0. cirzcta and T. minor alike. The latter species seems to be unable to actively influence the transpiration rate (VERHOEF and WITTEVEEN. 1980). Ach-nowlerlgements-The author is grateful to Drs. E. N. G. J~~SSE and N. D. DE WITH for helpful discussions, to Dr. J. M. SKELDING, Westfield College, University of London, for the introduction into the technique of measuring haemolymph osmolarity, to Mr. G. W. H. VAN DEN BERG for drawing the figures and to Miss. S. RICHTER for typing the manuscript.
759
REFERENCES BARTON BROWNE L. and VAN GERWEN A. C. M. (1976) Regulation of water ingestion by the locust, Chortoicetes terminifera: the effect of injections into the haemolymph. Phvsiol. Em. 1, 159-167. BEAMENTJ. W. L. (1976) The ecology of cuticle. In The Insect Integument (Ed. by HEPBURN H. R. ), pp. 359-374. Elsevier, Oxford. BERNAYS E. A. and CHAPMAN R. F. (1974) The effect of haemolymph osmotic pressure on the meal size of nymphs of Locusta migratoria L. J. exp. Biol. 61, 473480. BERRIDGE M. J. (1965) The physiology of excretion in the cotton stainer, Dysdercus fasciatus Signoret. II. Inorganic excretion and ionic regulation. J. exp. Biol. 43; 523-533. BERRIDGE M. J. (1970) Osmoregulation in terrestrial arthropods. In Chemical Zoology (Ed. by FLORKIN M. and SCHEERB. T.) Vol. V. part A, pp. 287-320. Academic Press, New York. BR~~CK E. (1976) Die Durchlsssigkeit der Cuticula von Blaberus trapezoideus und Periplaneta americana fiir eine wlssrige anorganische PhosphatlBsung (Blattaria: Blaberidae/Blattidae). Ent. Germ. 2, 320-327. DAVIS M. T. B. (1974) Critical temperature and changes in cuticular lipids in the rabbit tick, Haemaphysalis leporispalustris. J. Insect Physiol. 20, 1087-l 100. EDNEY E. B. (1977) Water Balance in Land Arthropods. Springer, Berlin. HADLEY N. F. (1977) Epicuticular lipids of the desert tenebrionid beetle, Eleodes armata: seasonal and acclimatory effects on composition. Insect Biochem. 7, 277-283. HALE W. G. (1965) Observations on the breeding biology of Collembola. Pedobiologia 5, 161-177. HOLDICH D. M. and MAYE_SK. R. (1976) Blood volume and total water content of the woodlouse, Oniscus asellus, in conditions of hydration and desiccation. J. Insect Ph.vsiol. 22, 547-553. JOOSSEE. N. G. and TFXERINK G. J. (1977) The role of food in the population dynamics of Orchesella cincta (Linnt) (Collembola). Oecologia (Berl.) 29, 189-204. J~IXSE E. N. G. and VERHOEF H. A. (1974) On the aggregational habits of surface dwelling Collembola. Pedobiologia 14, 245-249. LEE R. M. (1961) The variation of blood volume with age in the desert locust (Schistocerca gregaria Forsk.). J. Insect Physiol. 6, 36-5 1. NOBLE-NESBITTJ. (1963) Transpiration in Podura aquatica L. (Collembola, Isotomidae) and the wetting properties of its cuticle. J. exp. Biol. 40, 681-700. OKASHA A. Y. K. (1971) Water relations in an insect, Thermobia domestica. I. Water uptake from sub-saturated atmospheres as a means of volume regulation. J. e.xp. Biol. 55,435448. SUTTON S. L. (1972) Woodlice. Ginn, London. TREHERNEJ. E. and WILLMER P. G. (1975) Hormonal control of integumentary water-loss: evidence for a novel neuroendocrine system in an insect (Periplaneta americana). J. exp. Biol. 63, 143-159. VANNIER G. (1973) Etude de la transpiration chez un Insecte Collembole au tours de son exuviation. C.r. hebd. Skanc. Acad. Sci. Paris 211, 2231-2234. VANNIER G. and VERHOEFH. A. (1978). Effect of starvation on transpiration and water content in the populations of two coexisting Collembola species. Camp. Biochem. Physiol. 6OA, 483489. VERHOEF H. A. and WIT~EVEENJ. (1980) Water balance in Collembola and its relation to habitat selection; cuticular water loss and water uptake. J. Insect Physiol. 26,201-208. VERHOEFH. A. and NAGELKERKEC. J. (1977) Formation and ecological significance of aggregations in Collembola; an experimental study. Oecologia (Bed.) 31, 215-226. VERHOEFH. A., NAGELKERKEC. J. and JOOSSE E. N. G. (1977)
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Aggregation pheromones in Collembola. J. Insecf Physiol. 23, 1009-1013. WOODRINGJ. P. (1974) Effects of rapid and slow dehydration
on the haemolymph osmolarity and Na+-K+ concentration in the millipede Pachydesmus crassicutis. Camp. Biochem. Physiol. 49A, 115-I 19.