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Brazilian Journal of Medical and Biological Research

Print version ISSN 0100-879XOn-line version ISSN 1414-431X

Braz J Med Biol Res vol. 31 no. 8 Ribeirão Preto Aug. 1998 

Braz J Med Biol Res, August 1998, Volume 31(8) 1069-1073

Annual changes in serum calcium and inorganic phosphate levels and correlation with gonadal status of a freshwater murrel, Channa punctatus (Bloch)

S.K. Srivastav and A.K. Srivastav

Department of Zoology, University of Gorakhpur, Gorakhpur, India

down.gif (51 bytes) Abstract
Material and Methods
Correspondence and Footnotes


Adult Channa punctatus murrels of both sexes (60-80 g) were collected locally from Ramgarh Lake during the second week of every month (10 individuals of each sex/month) throughout the year. Blood samples were collected and analyzed for serum calcium and phosphate levels by the methods of Trinder (1960) and Fiske and Subbarow (1925), respectively. Gonads were fixed to judge the state of maturation of the fish. Males exhibited no change in serum calcium levels throughout the year in correlation with testicular maturation. However, serum phosphate levels exhibited a rise in correlation with the increased gonadosomatic index. Females showed marked seasonal changes in serum calcium and phosphate levels which were associated with ovarian maturation (vitellogenesis).

Key words: calcium, inorganic phosphate, reproduction, teleost, vitellogenesis


Vitellogenesis involves hepatic formation of yolk proteins (vitellogenin) and raises blood vitellogenin content followed by deposition of two forms of yolk in the ovary: yolk vesicles (mucopolysaccharides) and yolk granules (phospholipoproteins). Several authors have correlated the increased blood calcium content with ovarian maturation (1-7). Estradiol has been shown to stimulate hepatic formation of yolk proteins (vitellogenins) and to raise serum vitellogenin levels and the protein-bound fraction of plasma calcium levels (8-19). However, few studies describe the relationship between blood phosphate level and gonadal maturation in fishes (7,20). Oguri and Takada (goldfish; 11), Woodhead (arctic cod; 12) and Singh and Srivastav (catfish; 7) have found no correlation between serum calcium level and testicular maturation.

In the present study an attempt was made to investigate changes in serum calcium and phosphate levels in relation to ovarian maturation (vitellogenesis) and testicular maturation in a freshwater murrel, Channa punctatus.

Material and Methods

Ten male and ten female adult Channa punctatus murrels (15-20 cm; 60-80 g) were collected during the second week of every month throughout the year. Body and gonad weights were recorded for each animal to assess the variation in gonadosomatic index (GSI): GSI = gonad weight/body weight x 100.

Blood samples were collected under MS222 anesthesia by sectioning the caudal peduncle. Sera were separated by centrifugation and analyzed for calcium and phosphate levels according to the methods of Trinder (21) and Fiske and Subbarow (22), respectively. For histological studies gonads were fixed in Bouin's fixative. Sections were cut at 6 µm and processed with hematoxylin-eosin (HE).

To test the statistical significance, ANOVA, Student-Newman-Keuls and Dunnett multiple comparison tests were used.


The annual gonadal cycle of Channa punctatus can be divided into five phases, as shown in Table 1.


No change in serum calcium level was recorded throughout the year (Figure 1A) in relation to testicular cycle. However, serum phosphate level (Figure 1B) exhibited a rise corresponding to an increased GSI (Figure 1C).


The lowest serum calcium levels were recorded during the resting phase (March and October), with values increasing thereafter and peaking in June and early January (Figure 1A) when the end of the prespawning phase occurs for both reproductive peaks. During the spawning and postspawning phases, serum calcium levels showed a progressive fall (Figure 1A).

Serum phosphate levels increased with increasing GSI (Figure 1B and C). The serum calcium and phosphate levels of male and female Channa punctatus during different reproductive phases are shown in Figure 2A and B.

ANOVA indicated that the seasonal differences in serum calcium and phosphate values of males were not significant, whereas in females the serum calcium (P<0.03; F = 5.162) and phosphate values (P<0.05; F = 4.74) were significant. According to the Student-Newman-Keuls test, the serum calcium and phosphate levels of males and females showed no significant seasonal differences. According to the Dunnett multiple comparison test (considering resting phase as control), the serum calcium levels of preparatory, prespawning and spawning phases in females were significantly different from controls (P<0.05).

Figure 1 - Seasonal changes in serum calcium (A) and phosphate (B) levels and gonadosomatic index (C) of male and female Channa punctatus. Each value represents the mean ± SD of ten specimens.

[View larger version of this image (81 K GIF file)]

Figure 2 - Serum calcium (A) and phosphate (B) levels of male and female Channa punctatus during different reproductive phases. Each value represents the mean ± SD of ten specimens. RP = Resting phase; PP = preparatory phase; PS = prespawning phase; SP = spawning phase; PO = postspawning phase.

[View larger version of this image (50 K GIF file)]


In the present study, in agreement with reports by others, no correlation was found between serum calcium level and testicular maturation (6,7,9,11,20). In contrast to these reports, Woodhead and Woodhead (10) and Woodhead (12) have suggested a positive correlation between blood calcium level and testicular maturation in arctic cod and sea cod. Moreover, administration of testosterone propionate to male goldfish and killifish had no effect on blood calcium levels (23,24).

Bjornsson et al. (15) have reported that female rainbow trouts have higher levels of protein-bound calcium than males from September to February. Balbontin et al. (20), however, recorded fluctuations in the serum calcium levels of both sexes of hake (Merluccius gayi gayi) throughout the year and reported that calcium levels remained relatively constant from December to August in killifish. The authors suggested that this difference between male hake and killifish may correspond to different patterns of sex hormone secretion.

In male Channa punctatus, serum phosphate levels seemed to increase with increasing GSI even though the statistical results were not significant. A similar correlation has also been reported earlier by Balbontin et al. (20) for killifish and by Singh and Srivastav (7) for Heteropneustes fossilis.

A marked seasonal variation in serum calcium level was observed in female Channa punctatus, associated with ovarian maturation (i.e., different phases of vitellogenesis), in agreement with earlier observations (5-7,9-12,15,20,25). Bjornsson and Haux (26) reported that free plasma calcium levels are not affected, but the increase in total plasma calcium is due to the appearance of the calcium-containing yolk protein precursor vittellogenin in plasma.

Nagler et al. (27) have reported a linear relationship between increasing levels of indirect indicators such as serum total phosphoprotein phosphorus, alkali-labile phosphoprotein phosphorus, total calcium and increasing level of serum vitellogenin in mature female rainbow trout (Salmo gairdneri). The enhanced secretion of estrogen during the sexual maturation of females increases the calcium level (6,28). Bailey (23), Ho and Vanstone (8), Fleming et al. (9), Chan and Chester Jones (14) and Woodhead (13) detected increased serum calcium levels in female fish after the administration of estradiol. In female fish estradiol administration increases serum calcium level, as also observed in males of Carassius auratus (23), Fundulus kansae and Fundulus catenatus (9), Oncorhynchus nerka (8), Gadus morhua (13) and Fundulus heteroclitus (24). Serum calcium levels fell during the spawning and postspawning phases in Channa punctatus, in agreement with the earlier investigations of Fontaine et al. (25), Swarup et al. (6) and Singh and Srivastav (7).

We conclude that there is no correlation between the serum calcium levels of male Channa punctatus and testicular maturation; however, serum phosphate increased with increasing GSI. In females, the serum calcium and phosphate levels exhibited a progressive increase with ovarian maturation.


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Correspondence and Footnotes

Address for correspondence: S.K. Srivastav, Department of Zoology, University of Gorakhpur, Gorakhpur 273 009, India.

S.K. Srivastav is the recipient of a fellowship from CSIR, New Delhi. Received August 6, 1997. Accepted May 19, 1998.

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