HISTOARCHITECTURAL CHANGES IN THE ADRENAL GLAND OF THE FEMALE BAT TAPHOZOUS KACHHENSIS (DOBSON) DURING ESTRUS AND PREGNANCY

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1 Journal of Cell and Tissue Research Vol. 11(2) (2011) ISSN: (Available online at Original Article HISTOARCHITECTURAL CHANGES IN THE ADRENAL GLAND OF THE FEMALE BAT TAPHOZOUS KACHHENSIS (DOBSON) DURING ESTRUS AND PREGNANCY CHAVHAN, P. R., DHAMANI, A. A. AND MISAR, S. D. Department of Zoology, N. H. College, Bramhapuri E. mail: Received: June 19, 2011; July 25; 2011 Abstract: The present study shows the cyclical alternation in the activity of adrenal gland of the bat Taphozous kachhensis during different phases of the reproductive cycle. The adrenal gland is made up of steroidogenic and chromaffin cells. The gland during pregnancy is much larger than that of other phases of reproductive cycle. The cortex is completely encircles the medulla in Taphozous kachhensis. The medulla in this species consists of epitheloid cells arranged in smaller groups surrounded by blood capillaries. The zona glomerulosa consist of numerous bulbous structures which appear as acinus like group of cells. The zona fasciculata occurs in the form of irregular short cords; one or two cells thick arranged radially and merge with each other. These cords consist of large polyhedral vacuolated cells. The zona reticularis is also increase in size and measures. The cells are hypertrophied. Cytoplasm is eosinophilic with darkly stained nucleus and more vacuolated. Key words: Adrenal gland, Bat? INTRODUCTION Adrenal gland is a key organ in bat as it plays an important role in metabolism and homeostasis of animals. The life saver and life protecting hormone secreted by adrenal gland, helps to withstand the stress and trauma during pregnancy. Mammalian adrenal is unique among the vertebrate where the steroidogenesis and chromaffin cells are separated as cortex and medulla respectively. The light microscopic observations of adrenal gland are known in few species of bats viz., Eptrsicus fuscus and Anatrozous pallidus Anatrozous pallidus [1] Megaderma lyra lyra [2] Miniopterus schreibersii [3] Rousettus leschenauti and Pteropus gigantius [4] Cynopterus sphinx and Taphozous longimanus [5] Taphozous melanopogon [6] Hipposideros lankadiva [7] and Taphozous longimanus [8]. The study of weight, size and shape of right and left adrenal gland in pregnant and nonpregnant bats show the significant difference in T. melanopogon [9] The weight of adrenal gland is more during pregnancy than the nonpregnant condition observed in Rousettus aegyptiacus [10]. The objective of present study is to analyse the histoarchitectural changes in the adrenal gland of the female bat Taphozous kachhensis (dobson) during estrus and pregnancy. MATERIALS AND METHODS Taphozous kachhensis (Dobson) is an exclusive Indian Emballonuride bat found in caves, tunnels and temples. The bat is selected for present study to because of its unique habits. The gestation length of adult female of the species Taphozous kachhensis (Dobson) is about 100 days. The collection of the specimens commenced in February 2006 and the last specimen for the present study was collected in May 2009.The specimen of Taphozous kachhensis were collected from Ambai Nimbai, 45 kilometers from Bramhapuri in Maharashtra. Many collections were made during the breeding season so as to coincide 2857

2 J. Cell Tissue Research PLATE 1 Fig. 1: Transverse section cortex (CX) and medulla (MD). Note the oblong shape of adrenal gland. X 40 Fig. 3 Fig. 2: Transverse section zona glomeru-losa (ZG), zona fascic-ulata (ZF), zona reticul-aries (ZR) and medulla (MD). X 1000 Fig. 1 Fig. 3; Transverse section vacuolated cells of zona glomerulosa (ZG) with darkly stained spherical nuclei. X 1000 Fig. 2 Fig. 4 Fig. 5 Fig. 6 Fig. 4: Transverse section elongated cell cords of zona fasciculata (ZF) with vacuolated cell cytoplasm. X 1000 Fig. 5: Transverse section the cell of zona reticularis (ZR) are in the form anastomosing cords and having varying degree of shape and size. Each cell has eosinophilic cytoplasm with vesicular nucleus. Blood spaces are observed in the network of cell cords of zona reticularies. X 1000 Fig. 6: Transverse section medulla(md) consist of cells arranged in irregular strands or short cords surrounded by blood capillaries. Cells contain darkly stained vesicular nucle-us. Cytoplasm of cell is basophilic and granular and in some cells Vaculation is observed. X

3 Chavhan et al. with the time of reproductive cycle and to get an accurate pregnancy record. During day time their roosting places were visited and the specimens were netted at random with the help of a butterfly net. These bats are very sluggish in nature. After collection they were sexed and only the females were brought to the laboratory. Weight recorded with sensitive balance before they were sacrificed. The Adrenal glands were dissected out and fixed in alcoholic Bouin s fluid. After fixation for 24 hrs tissue were washed with 70% ethanol. For histological observation, the tissues were later dehydrated in various grades of alcohol, cleared in xylol, embedded in paraffin wax. The tissues were cut at 5 to 6 mµ with the help of Leica 2417 microtome. The sections were stained with haematoxylin and eosin for routine histological examination. RESULTS The adrenal gland of Taphozous. kachhensis is situated retroperitoneal on each side occupying the antero-mesialposition abutting against the ipsilateral kidney to which it is firmly attached by renal fascial and is enclosed by a thick capsule. The adrenal gland can be distinguished as an outer cortex and inner medulla, even though a distinct circum medullary connective tissue capsule separating the two regions is missing. The medulla is relatively extensive in this species of bat. The cortex completely encircles the medulla intaphozous kachhensis. Histology: The histological changes in the adrenal gland of the Taphozous kachhensis during different phases of reproductive cycle are presented in figure (see photo plates 1,2,3). A. ADRENAL GLAND DURING ESTRUS PERIOD (Photo plate 1): Adrenal gland of Taphozous kachhensis is round to oval in shape and encloses in a connective capsule made up of spindle fibrous cells (Fig.1). Thin strand of connective tissue or trabeculae extend from capsule between the columns of cells in the cortex. Beneath the capsule, glands thave two distinct zonations the cortex and medulla. On the basis of the observations on histology of the steroidogenic cells, cortex is further divided in to three zones namely zona glomerulosa, zona fasciculata and zona reticularies. Zona glomerulosa: This is the smallest zone of adrenal cortex measuring 90 µ during the estrous period. This zone consists of small spherical cells that are compactly arranged and appear acinus like group of cells. These cells have lightly stained cytoplasm with small nuclei with clear nucleolus. Chromatin clumps are observed. Cytoplasm is eosinophilic and granular. Vaculation is observed in the cytoplasm of some cells. Acinar membrane is well defined (Fig 2). Zona fasciculata: This is the widest cortical zone observed in the adrenal cortex measuring 300 µ during estrous period. Zona fasciculata consist of large polyhedral or cuboidal cells arranged in cords. Cytoplasm is eosinophilic and vacuolated due to the presence of lipid droplets. Nucleus is darkly stained. Zona fasciculata merges with zona glomerulosa above and zona reticularies below. The cords are separated from each other by connective tissue and blood capillaries (Fig. 3). Zona reticularis: This zone is present just below the Zona fasciculata and measures about 90 µ during estrous period. The cell of Zona reticularis are in the form of anastomosing cords and having varying degree of shape and size. Each cell has eosinophilic cytoplasm with vesicular nucleus. Blood spaces are observed in the network of cell cords of zona reticularies (Fig. 4). Medulla: Medulla is clearly demarcated from the cortex in Taphozous kachhensis. It measures about 270 µ during the estrous period of the reproductive cycle. Medulla consists of cells arranged in irregular strands or short cords surrounded by blood capillaries. Cells contain darkly stained vesicular nucleus. Cytoplasm of cell is basophilic and granular. In some cells Vaculation is observed (Fig 5). ADRENAL GLAND DURING EARLY PREGN- ANCY (Photo plate 2): Adrenal gland of Taphozous kachhensis is oval to elongated in shape. Both cortex and medulla are well developed in this animal. Zona glomerulosa: Zona glomerulosa increase in size and measure about 120µ. Cells of glomerulosa are elongated with darkly stained nucleus. Cytoplasm is eosinophilic and Vaculations are observed in cytoplasm (Fig. 1). 2859

4 J. Cell Tissue Research Fig. 1 Fig. 2 PLATE 2 Fig. 3 Fig. 4 Fig. 5 Fig. 1: Transverse showing cortex (CX) and medulla (MD). Note the triangular shape of adrenal gland. X 40 Fig. 2: Transverse showing zona glomerulosa (ZG), zona fasciculata (ZF), zona reticularies (ZR) and medulla (MD). X 1000 Fig. 3; Transverse showing elongated cells of zona glomerulosa (ZG) with darkly stained nuclei and vacuolation in cytoplasm. X 1000 Fig.4; Transverse showing cells of zona fasciculata (ZF). cells are hypertrophied and nucleus is vesicular, round and darkly stained. The cytoplasm is eosinophilic and more vacuolated. X 1000 Fig. 5: Transverse showing the cell of zona reticularis (ZR). Cell cytoplasm is eosinophilic with darkly stained nucleus and more vacuolated as compare to estrus. X 1000 Fig. 6: Transverse early pregnancy showing well developed medulla (MD) consists of cells arranged in acini surrounded by blood capillaries. X 1000 Fig

5 Chavhan et al. Zona fasciculata: During early pregnancy Zona fasciculata is smaller in size as compared to estrus period of reproductive cycle and measure about 113 µ. The cells are hypertrophied and nucleus is vesicular, round and darkly stained. The cytoplasm is eosinophilic and more vacuolated as compared to estrus because of high lipid droplets (Fig. 2). Zona reticularis: Zona reticularis increases in size and measures 105 µ. The cells are hypertrophied. Cytoplasm is eosinophilic with darkly stained nucleus and more vacuolated as compared to estrus. Zona fasciculata merge with Zona reticularies without any clear demarcation (Fig. 3). Medulla: Medulla is more developed during early pregnancy and measures about 580 µ. Hypertrophied medullary cells are arranged in the acini encircling the blood spaces. Cell cytoplasm is lightly stained and granular with vaculation. Blood supply is also increased and blood vessels are observed encircling the cells (Fig 4). ADRENAL GLAND DURING LATE PREGN- ANCY (Photo plate 3): Adrenal gland of Taphozous kachhensis is oval to elongated in shape. Cortex is reduced in size and medulla extensively developed (Fig. 1). Zona glomerulosa: Zona glomerulosa is made up of acinar structure. Vaculation is more pronounced in the cytoplasm. Zona glomerulosa measure about 105 µ. Cell cytoplasm is lightly stained with round nucleus (Fig. 2). Zona fasciculata: Zona fasciculata increases in diameter and measure about 130 µ. Zona fasciculata merge with zona glomerulosa above. It is in the form of cords which are not radially arranged and irregular in distribution. Cell cytoplasm is lightly stained. Cytoplasmic vacuolation is more pronounced (Fig. 3). Zona reticularis: Zona reticularis is measure about 95µ in diameter. Cells are large with faintly stained nucleus observed in the center of the cell and shows prominent nucleolus. Lipid vacuoles are seen in the cells (Fig. 4). Medulla: Medulla is very large and measures about 600 µ in diameter. It is made up of a group of cells and surrounded by blood capillaries. The compactly arranged group of cells are seen scattered. Cell cytoplasm appears granular and vacuolated (Fig. 5). DISCUSSION In Taphozous kachhensis, the size and weight of left adrenal gland is always higher than the right adrenal gland during different phases of reproductive cycle. Similar observations are reported in T. melanopogon [6]. The weight of the adrenal gland increases during estrus and is maximum during early pregnancy. It decreases at mid-pregnancy. However, it again increases at late pregnancy. The cortex and medulla of adrenal gland of bat, Taphozous kachhensis are clearly marked. Similar observations were made in the bat, M. lyra lyra [2], M. schreibersii [3], and R. leschenaulti [4]. In Taphozous longimanus [8] only two zone are distinct. The adrenal cortex is distinct in three zones viz., zona glomerulosa, zona fasciculata and zona reticularis. A distinct zonation of the cortex is observed in M. lyra lyra [2], M. schreibersii [3], V. pipistrellus [11]. The adrenal gland of Taphozous kachhensis is oval to oblong in shape and is enclosed in the connective tissue capsule. Beneath the capsule lies a small cortical zone, the zona glomerulosa. It consists of numerous acinus like group of cells. Similar structure is also reported in the zona glomerulosa of H. lankadiva [7]. However, the cells are arranged in columns in R. leschenauti [4]. The glomerulosa merges with the zona fasciculata. The zona fasciculata occurs in the form of irregular short cords; one or two cells thick arranged radially and merge with each other. These cords consist of large polyhedral vacuolated cells. Similar pattern of arrangement in zona fasciculata is observed in Rousettus leschenauti [4]. These cords are made up of two cells thick polyhedral cells in P. gigantius [4], M. lyra lyra [2].The zona reticularies is in the form of anastomosing cords of cells, which merge with zona fasciculata aboveand below medulla. However, zona reticularies is absent in T. longimanus [8]. The medulla of T. kachhensis is well developed. Cells are arranged in irregular strands or short cords surrounded by blood capillaries. Similar structure of medulla is reported in T. longimanus [8], R. leschenauti and P. gigantius [4]. Reported the presence of two types of medullary cells in Pteropus and Rousettus on the basis of staining with basic dyes. Adrenal gland during estrus shows significant 2861

6 J. Cell Tissue Research PLATE 3 Fig. 1: Transverse showing cortex (CX) and well developed medulla (MD). Note the oval shape of adrenal gland. X 40 Fig. 3 Fig. 2: Transverse showing zona glomerulosa (ZG), zona fasciculata (ZF), zona reticularies (ZR) and medulla (MD). X 400 Fig. 1 Fig. 3; Transverse showing elongated cells of zona glomerulosa (ZG) with darkly stained spherical elongated nuclei. X 400 Fig. 4 Fig. 5 Fig.4: Transverse showing cells of zona fascic-ulata (ZF) with large vacuolated cell cords and lightly stained nuclei. X 400 Fig. 5: Transverse showing the large cells of zona reticularis (ZR). Note the spherical shape of nucleus with prominent nucleoli. X 400 Fig. 2 Fig. 6: Transverse showing well developed med-ulla (MD). cells are scattered and surrounded by blood capillaries. X 400 Fig

7 Chavhan et al. difference at structural level. During estrus the zona fasciculata is well developed and made up of polygonal cells with vesicular nuclei and vaculations are more pronounced due to lipid droplets. Thus zona fasciculata is more developed than two other zones. Such distinct features are also observed in Taphozous melanopogon [6] and Hipposideros lankadiva [7] during the non pregnant state. There is an increase in the size pregnancy, it is oval to elongate in form. Histoarchitecture of adrenal gland is advanced over that of adrenal gland of estrus bat. The cells of glomerulosa are elongated. Cytoplasm shows eosinophilic granules and vacuoles are more in the cytoplasm. During pregnancy, the zona fasciculata is well developed than the zona glomerulosa. The medulla is well developed in the adrenal glands in pregnant bat. Seasonal changes in adrenal gland in female bat of T. melanopogon during pregnancy and non-pregnancy have been studied [6]. Thus adrenal gland of these bats show significant differences of structure over adrenal gland of non pregnant bats supporting present observations of Taphozous kachhensis during non pregnancy and pregnancy. in Hipposideros lankadiva (Kelaart), Ph. D. thesis, RTM NagpurUniversity, Nagpur (2004). [8] Nerkar, A.A. : Electron Microscopic Studies on the Endocrine gland and Reproductive organs of Emballonuride female bat, Taphozous longimanus (Hardwicke) during Reproductive Cycle. Ph.D thesis R.T.M. Nagpur uni. Nagpur (2007). [9] Lowry, M.L. and Lall, S.B.: Evidence for the involment of the adrenal gland is gestation junction of Taphozous melanopogon melanopogon (chiroptera) displaying absolute dextral dominance of genital tract. European Bat Research, Charles Uni. Press, Praha (1987). [10] Baranga, J.: The weight changes of tropical fruit bat, Rousettus aegyptiacus E. Geoffory. Sounerdruck aus Zeitschrift feir Saugetierkunde 45: (1980). [11] Saidapur, S.K. and Nadkarni, V.B.: Endocrinology, 67: 1105 (1976). ACKNOWLEDGEMENTS I am thankful to Dr N S Kokode for providing us necessary facility for the laboratory work during the course of this work. REFERENCES [1] Gorbman, A. and Bern, H.A.: A Textbook of Comparative Endocrinology. Wiley, New York (1962). [2] Bhima Rao, B.S. and Devaraj Sarkar, H.B.: Cur. Sci., 44: (1975). [3] Planel, H., Guilhem, A. and Soleihavoup, J.P.: Comptes Rendus de I Association des Anatomistes, 47: (1961). [4] Sapkal, V.M.: Curr. Sci., 4: (1978). [5] Karim, K.B., Gopalkrishna,A. and Gadehok, H.: Current Sci., 13: ( 1979). [6] Lowry, M.L. and Lall, S.B.: Morphometric and Enzymological changes in Melanopogon melanopogon (Trmmic) from the Multiparus and parous state. Seventh international bat research conference. University of Abrod, U.K. pp (1986). [7] Seraphim, E.R.: Endocrine Interaction during different phases of the Female Reproductive Cycle 2863