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Pineal-adrenal relationship in a lizard Calotes versicolor
GENERAL
AND
COMPARATIVE
ENDOCRINOLOGY
Pineal-Adrenal
39, 414-416 (1979)
Relationship
in a Lizard Calotes
versicolor
Pinealectomy caused hypertrophy and melatonin atrophy of the adrenal cortex as reflected by alterations in the percentage of total adrenal volume and weight of the adrenal in the lizard, Calotes versicolor.
Attempts to establish a relationship beAll animals were kept in a room fully exposed to natural day length and temperatween pineal and adrenal have generally yielded consistent results (Kappers , 197 1). ture. After 30 days of the treatment animals Following pinealectomy, increase in ad- were killed and the adrenals carefully removed, cleaned, weighed and fixed in renal hormone level has been noted in cats, rabbits, and sheep by Kitay and Altschule Bouin-Hollande’s fluid, section at 5 pm, (1964), and more critically in rats and mice and stained with Weigert’s hematoxylin, erythrocin, and counterstained with light by Ogle and Kitay (1976), Dickson (1971), green. The percentage of total adrenal volGiordano et al. (1970), and Vaughan et al. ume was determined by Camera Lucida (1972) who measured aldosterone and corticosterone levels. Contrary results sug- diagrams of each serial section drawn on graph paper (Miller and Riddle, 1942). gesting elevated ACTH level by pineal Calotes versicolor has a pair of adrenals. extracts and decreased level by pinealectomy have also been reported (Cassano et These adhere to the anterio-inner end of the vas deferens and cover the outer margin of al., 1971; Dickson, 1971). It has also been suggested that pineal acts directly on ad- the testis. The right adrenal is slightly longer than the left. Both adrenals lie anrenal cortex leading to reduced production of 4,3-ketonic corticosteroids (Giordano ef terior to the cephalic end of the kidneys and and yellowish in color. al. 1970). In lower vertebrates, excepting a are nonpigmented The histology of the gland confirms to the suggestion of the pineal adrenal axis (Kappattern given in Gabe (1970) and Lofts pers, 1971), there is practically complete (1978). lack of information on the pineal-adrenal The adrenal gland of saline injected anirelationship. Experiments were therefore planned to determine whether pineal has an mals had well-defined cortical cords and weighed significantly less than the effect on the adrenal of a lizard. pinealectomized saline treated animals. Male garden lizards, CaIotes versicolor, tissue ratio of the intact collected during June were used on this CorticaVchromaftin study. Forty eight animals, after 15 days of saline injected animals was also significantly less than pinealectomized saline acclimatization to captivity, were divided into six groups of 8 lizards each. Lizards of treated ones (Table 1). There was no significant difference in group I were sham operated and of group II pinealectomized (Haldar, 1976). Animals of either weight or in any of the parameters studied between the 50 pg melatonin and other groups were treated with different the saline-injected intact animals (Table 1). doses of melatonin (Sigma) in saline and of I and II with normal saline (0.9%; Table 1). But in 100, 200, and 400 fig melatonin injected lizards significant decrease had ocThe total dose of the hormone was injected in 30 daily ip injections. The volume of curred in weight, area of vascularization, saline or of hormone injected at a time was and cortical cord width of the adrenals (Table 1). Between the three higher doses 0.1 ml. only. 414 00116480/79/110414-03$01.ooM) Copyright All ii&s
@ 1979 by Academic Press. Inc. of repmduciioo in any form -cd.
415
NOTES TABLE 1 CHANGES IN ADRENAL ACTIVITY IN PINEALECTOMIZED AND MELATONIN-TREATED &dotes
versicofor
Percentage of total adrenal volume + SEM Experimental condition
No. of animals
Adrenal wt.O (rng ” SEM)
cortical tissue
Group I Sham-operated + saline
8
14.1** 20.5
63.8A 52.1
31.2c kO.6
5.0 20.1
1:2*
Group II Pinealectomized + saline
8
19.2s.’ -cl.4
76.2* 26.6
19.0** k-o.4
5.0 21.3
1:4**
Group III Normal + 50 @g melatonin
8
14.4A LO.5
60.26 25.0
37.2c 23.7
2.2c*** 20.1
1:2B
Group IV Normal + 100 Fg melatonin
8
12.Y kO.8
50.7c* -cl.4
48.3c* *1.3
2 4c*** r0.2
l:l’*
Group V Normal + 200 @g melatonin
8
11.9c* 20.4
42.7c** ~2.8
56. lc** 22.8
2.6c*** kO.1
1:1c**
Group VI Normal + 400 pg melatonin
8
11.5c* +0.3
33 2c*** k-1.3
36.Y*** + 1.3
2.3c*** kO.1
1.1c***
Chromafin
Vascular area
I/C ratio
” Calculated as mg/lOO g body wt. * Significance of difference from pinealectomized + saline A < 0.05, B < 0.01, C c 0.001. p Significance of difference from sham-operated + saline * < 0.05, ** < 0.01, *** < 0.001.
however there was no significant difference in any of the parameters examined (Table 1). Pinealectomy led to hypertrophy and melatonin to atrophy of the adrenal cortex as reflected by percentage of total adrenal volume and weight in Calotes versicolor. Similar results of pinealectomy have been described in mammals (Kinsen and Singer, 1967; Kinsen et al., 1967). Pinealectomy was found to increase both aldosterone and corticosterone levels (Karppanen et al. 1970). Wurtman et al. (1959) have also reported adrenal enlargement in rats after pinealectomy and atrophy following pineal extract administration. Present results are similar to those reported in rats (Wurtman et al., 1959; Anton-tay, 1971; Vaughan et a[. , 1972) and indicate that in reptiles (also) the pineal-adrenal relationship is of the mammal type, and that melatonin possibly represents the pineal hormone. The pineal may influence the adrenal
either by way of the hypothalamohypophyseal complex or directly. In either case it seems to produce a check on the activity of the adrenal. In Calotes this effect would be prominent especially during winter when days are short and pineal maximally active. But, how adrenal activity is reduced during winter, when the animals normally should be under considerable stress is not known. Could it be that winter sleep reduces the adrenal response of the animals? Again, what is the role, if any, of corticoids in reptilian physiology during winter sleep? Answers to some of these questions will help us to understand better the physiology of reptiles in relation to the pineal. REFERENCES Anton-tay, F. (1971). Pineal-brain relationship. In Ciba Foundation Symposium: The Pineal Gland (G. E. W. Wolstenholme and A. Knight, eds.), pp. 213-227, Churchill Livingstone, London.
416
NOTES
Cassano, C., Torscu, A., Peruzy, A. DC. De., and Martin, C. (1971). Studi sul’ epilis indagin mell‘ animale de experiments e nell’ unomo. Folia Endocrinol. 14, 755-790. Chaubey, B. J. (1971). “Natural Biology and Endocrinology of the Indian Garden Lizard, Calotes versicolor (Daud) Male.” Doctoral dissertation, Banaras Hindu University, India. Dickson, K. L. (1971). Effects of the pineal gland in adrenalectomized rats. Acra Endocrinol. (KM) 70,438-444. Gabe, M. (1970). The adrenal. In “Biology of Reptilia” (C. Cart and P. Thomas, eds.), vol. 3, pp. 263-318. Academic Press, London/New York. Giordano, G., Balestri, R., Jacopino, E., Foppiani, E.. and Bartolin, S. (1970). Melatonin and steroidogenesis. Sunchalica. Arch. Maregliano Pat&. Clin. 25, 109-117. HaIdar, C. (1976). “Pineal in Reptiles: Structure and Function.” Doctoral dissertation, Banaras Hindu University, India. Kappers. J. A. (1971). 1tz “Ciba Foundation Symposium: The Pineal Organ: An introduction” (G. E. W. Wolstenholme and J. Knight, eds.) pp. 3-34. Churchill, London. Karppanen, H., Vapaatalo, H., Lahovaaru. S., and Paasmen, M. K. (1970). Studies with pinealectomized rat. Pharma 3, 76-84. Kinsen, G. A., and Singer, B. (1967). The pineal gland and the adrenal response to sodium deficiency in the rat. Neuroendocrinology 2, 283-293. Kinsen, G. A., Wahid, A. K., and Singer, B. (1967). Effect of chronic pinealectomy on adrenocortical hormone secretion in normal and hypertensive rats. Gen. Comp. Endocrinol 8, 445-454.
Kitay, J. I. and Altschule, M. D. (1964). “The Pineal Gland. Harvard Univ. Press, Cambridge, Mass. Lofts, B. (1978). “General Comparative and Clinical Endocrinology of Adrenal Cortex. Vol. 2 (I. Chester-Jones and I. W. Henderson, eds.). Academic Press, London/New York/San Francisco. Miller, R. A., and Riddle, I?. (1942). The cytology of the adrenal cortex of normal pigeons in experimentally induced atrophy and hypertrophy. Amer. .I. Anat. 71, 311-385. Ogle, T. F., and Kitay, J. I. (1976). Effect of pinealectomy on adrenal function in LGvo, in vitro in female rats. Endocrinology 98(l), 20. Vaughan, M. K., Reiter, R. J., and Vaughan, G. M. (1972). Effect of melatonin and other pineal indole in adrenal enlargement produced in male and female mice by pinealectomy, unilateral adrenalectomy , castration and cold stress. Neuroendocrinoiogy 10, 139- 154. Wurtman, R. J., Atschule, M. D., and Holmgren. U. (1959). Effects of pinealectomy and of a bovine pineal extract in rats. Amer. J. Physiol. 197, 108- 110. C. MISRA NEE HALDAR J. P. THAPLIYAL Department of Zoology Banaras Hindu University Varanasi 221005 India Accepted June 12. 1979
AND
COMPARATIVE
ENDOCRINOLOGY
Pineal-Adrenal
39, 414-416 (1979)
Relationship
in a Lizard Calotes
versicolor
Pinealectomy caused hypertrophy and melatonin atrophy of the adrenal cortex as reflected by alterations in the percentage of total adrenal volume and weight of the adrenal in the lizard, Calotes versicolor.
Attempts to establish a relationship beAll animals were kept in a room fully exposed to natural day length and temperatween pineal and adrenal have generally yielded consistent results (Kappers , 197 1). ture. After 30 days of the treatment animals Following pinealectomy, increase in ad- were killed and the adrenals carefully removed, cleaned, weighed and fixed in renal hormone level has been noted in cats, rabbits, and sheep by Kitay and Altschule Bouin-Hollande’s fluid, section at 5 pm, (1964), and more critically in rats and mice and stained with Weigert’s hematoxylin, erythrocin, and counterstained with light by Ogle and Kitay (1976), Dickson (1971), green. The percentage of total adrenal volGiordano et al. (1970), and Vaughan et al. ume was determined by Camera Lucida (1972) who measured aldosterone and corticosterone levels. Contrary results sug- diagrams of each serial section drawn on graph paper (Miller and Riddle, 1942). gesting elevated ACTH level by pineal Calotes versicolor has a pair of adrenals. extracts and decreased level by pinealectomy have also been reported (Cassano et These adhere to the anterio-inner end of the vas deferens and cover the outer margin of al., 1971; Dickson, 1971). It has also been suggested that pineal acts directly on ad- the testis. The right adrenal is slightly longer than the left. Both adrenals lie anrenal cortex leading to reduced production of 4,3-ketonic corticosteroids (Giordano ef terior to the cephalic end of the kidneys and and yellowish in color. al. 1970). In lower vertebrates, excepting a are nonpigmented The histology of the gland confirms to the suggestion of the pineal adrenal axis (Kappattern given in Gabe (1970) and Lofts pers, 1971), there is practically complete (1978). lack of information on the pineal-adrenal The adrenal gland of saline injected anirelationship. Experiments were therefore planned to determine whether pineal has an mals had well-defined cortical cords and weighed significantly less than the effect on the adrenal of a lizard. pinealectomized saline treated animals. Male garden lizards, CaIotes versicolor, tissue ratio of the intact collected during June were used on this CorticaVchromaftin study. Forty eight animals, after 15 days of saline injected animals was also significantly less than pinealectomized saline acclimatization to captivity, were divided into six groups of 8 lizards each. Lizards of treated ones (Table 1). There was no significant difference in group I were sham operated and of group II pinealectomized (Haldar, 1976). Animals of either weight or in any of the parameters studied between the 50 pg melatonin and other groups were treated with different the saline-injected intact animals (Table 1). doses of melatonin (Sigma) in saline and of I and II with normal saline (0.9%; Table 1). But in 100, 200, and 400 fig melatonin injected lizards significant decrease had ocThe total dose of the hormone was injected in 30 daily ip injections. The volume of curred in weight, area of vascularization, saline or of hormone injected at a time was and cortical cord width of the adrenals (Table 1). Between the three higher doses 0.1 ml. only. 414 00116480/79/110414-03$01.ooM) Copyright All ii&s
@ 1979 by Academic Press. Inc. of repmduciioo in any form -cd.
415
NOTES TABLE 1 CHANGES IN ADRENAL ACTIVITY IN PINEALECTOMIZED AND MELATONIN-TREATED &dotes
versicofor
Percentage of total adrenal volume + SEM Experimental condition
No. of animals
Adrenal wt.O (rng ” SEM)
cortical tissue
Group I Sham-operated + saline
8
14.1** 20.5
63.8A 52.1
31.2c kO.6
5.0 20.1
1:2*
Group II Pinealectomized + saline
8
19.2s.’ -cl.4
76.2* 26.6
19.0** k-o.4
5.0 21.3
1:4**
Group III Normal + 50 @g melatonin
8
14.4A LO.5
60.26 25.0
37.2c 23.7
2.2c*** 20.1
1:2B
Group IV Normal + 100 Fg melatonin
8
12.Y kO.8
50.7c* -cl.4
48.3c* *1.3
2 4c*** r0.2
l:l’*
Group V Normal + 200 @g melatonin
8
11.9c* 20.4
42.7c** ~2.8
56. lc** 22.8
2.6c*** kO.1
1:1c**
Group VI Normal + 400 pg melatonin
8
11.5c* +0.3
33 2c*** k-1.3
36.Y*** + 1.3
2.3c*** kO.1
1.1c***
Chromafin
Vascular area
I/C ratio
” Calculated as mg/lOO g body wt. * Significance of difference from pinealectomized + saline A < 0.05, B < 0.01, C c 0.001. p Significance of difference from sham-operated + saline * < 0.05, ** < 0.01, *** < 0.001.
however there was no significant difference in any of the parameters examined (Table 1). Pinealectomy led to hypertrophy and melatonin to atrophy of the adrenal cortex as reflected by percentage of total adrenal volume and weight in Calotes versicolor. Similar results of pinealectomy have been described in mammals (Kinsen and Singer, 1967; Kinsen et al., 1967). Pinealectomy was found to increase both aldosterone and corticosterone levels (Karppanen et al. 1970). Wurtman et al. (1959) have also reported adrenal enlargement in rats after pinealectomy and atrophy following pineal extract administration. Present results are similar to those reported in rats (Wurtman et al., 1959; Anton-tay, 1971; Vaughan et a[. , 1972) and indicate that in reptiles (also) the pineal-adrenal relationship is of the mammal type, and that melatonin possibly represents the pineal hormone. The pineal may influence the adrenal
either by way of the hypothalamohypophyseal complex or directly. In either case it seems to produce a check on the activity of the adrenal. In Calotes this effect would be prominent especially during winter when days are short and pineal maximally active. But, how adrenal activity is reduced during winter, when the animals normally should be under considerable stress is not known. Could it be that winter sleep reduces the adrenal response of the animals? Again, what is the role, if any, of corticoids in reptilian physiology during winter sleep? Answers to some of these questions will help us to understand better the physiology of reptiles in relation to the pineal. REFERENCES Anton-tay, F. (1971). Pineal-brain relationship. In Ciba Foundation Symposium: The Pineal Gland (G. E. W. Wolstenholme and A. Knight, eds.), pp. 213-227, Churchill Livingstone, London.
416
NOTES
Cassano, C., Torscu, A., Peruzy, A. DC. De., and Martin, C. (1971). Studi sul’ epilis indagin mell‘ animale de experiments e nell’ unomo. Folia Endocrinol. 14, 755-790. Chaubey, B. J. (1971). “Natural Biology and Endocrinology of the Indian Garden Lizard, Calotes versicolor (Daud) Male.” Doctoral dissertation, Banaras Hindu University, India. Dickson, K. L. (1971). Effects of the pineal gland in adrenalectomized rats. Acra Endocrinol. (KM) 70,438-444. Gabe, M. (1970). The adrenal. In “Biology of Reptilia” (C. Cart and P. Thomas, eds.), vol. 3, pp. 263-318. Academic Press, London/New York. Giordano, G., Balestri, R., Jacopino, E., Foppiani, E.. and Bartolin, S. (1970). Melatonin and steroidogenesis. Sunchalica. Arch. Maregliano Pat&. Clin. 25, 109-117. HaIdar, C. (1976). “Pineal in Reptiles: Structure and Function.” Doctoral dissertation, Banaras Hindu University, India. Kappers. J. A. (1971). 1tz “Ciba Foundation Symposium: The Pineal Organ: An introduction” (G. E. W. Wolstenholme and J. Knight, eds.) pp. 3-34. Churchill, London. Karppanen, H., Vapaatalo, H., Lahovaaru. S., and Paasmen, M. K. (1970). Studies with pinealectomized rat. Pharma 3, 76-84. Kinsen, G. A., and Singer, B. (1967). The pineal gland and the adrenal response to sodium deficiency in the rat. Neuroendocrinology 2, 283-293. Kinsen, G. A., Wahid, A. K., and Singer, B. (1967). Effect of chronic pinealectomy on adrenocortical hormone secretion in normal and hypertensive rats. Gen. Comp. Endocrinol 8, 445-454.
Kitay, J. I. and Altschule, M. D. (1964). “The Pineal Gland. Harvard Univ. Press, Cambridge, Mass. Lofts, B. (1978). “General Comparative and Clinical Endocrinology of Adrenal Cortex. Vol. 2 (I. Chester-Jones and I. W. Henderson, eds.). Academic Press, London/New York/San Francisco. Miller, R. A., and Riddle, I?. (1942). The cytology of the adrenal cortex of normal pigeons in experimentally induced atrophy and hypertrophy. Amer. .I. Anat. 71, 311-385. Ogle, T. F., and Kitay, J. I. (1976). Effect of pinealectomy on adrenal function in LGvo, in vitro in female rats. Endocrinology 98(l), 20. Vaughan, M. K., Reiter, R. J., and Vaughan, G. M. (1972). Effect of melatonin and other pineal indole in adrenal enlargement produced in male and female mice by pinealectomy, unilateral adrenalectomy , castration and cold stress. Neuroendocrinoiogy 10, 139- 154. Wurtman, R. J., Atschule, M. D., and Holmgren. U. (1959). Effects of pinealectomy and of a bovine pineal extract in rats. Amer. J. Physiol. 197, 108- 110. C. MISRA NEE HALDAR J. P. THAPLIYAL Department of Zoology Banaras Hindu University Varanasi 221005 India Accepted June 12. 1979