Research Article

 

Effect of Insulin on Post-AI Circulating Estrodiol-17β and LH in Crossbred Cattle

 

Mridula Sharma*, H.P.Gupta, Shiv Prasad

Department of Veterinary, Gynaecology and Obstetrics, College of Veterinary and Animal Sciences, G. B. Pant University of Ag. & Tech., Pantnagar-263145 Distt- U.S. Nagar, Uttarakhand.

 

Editor | Asghar Ali Kamboh, Sindh Agriculture University, Tandojam, Pakistan.

Received | May 22, 2018; Accepted | October 11, 2018; Published | November 06, 2018

*Correspondence | Mridula Sharma, Department of Veterinary, Gynaecology and Obstetrics, College of Veterinary and Animal Sciences, G. B. Pant University of Ag. & Tech., Pantnagar-263145 Distt- U.S. Nagar, Uttarakhand; Email: [email protected]

Citation | Sharma M, Gupta HP, Prasad S (2018). Effect of Insulin on post-AI circulating estrodiol-17β and LH in crossbred cattle. J. Anim. Health. Prod. 6(4): 103-107.

DOI | http://dx.doi.org/10.17582/journal.jahp/2018/6.4.103.107

ISSN (Online) | 2308-2801; ISSN (Print) | 2309-3331

Copyright © 2018 Sharma et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

 

Introduction

 

Application of metabolic hormones i.e. somatotrophin, insulin and insulin like growth factors (IGF-1) in regulation of ovarian functions in livestock is fairly a recent development (Purkayastha et al., 2015). Insulin and IGF-I either alone or in combination with gonadotrophins have been found to have a profound effect on steroidogenesis of cultured bovine granulosa cells (Singh et al., 2010, Gong et al., 1993b; Spicer et al., 1993). Garzo and Darrington (1984), Poretsky and Piper (1994) and Willis et al. (1996) suggested that acting at the ovarian level, insulin appears to potentiate the steroidogenic response to gonadotropins, both in vitro (Baithalu et al., 2013) and in vivo. In granulosa cells, this effect may be mediated by an increase in LH (Luteinising Hormone) receptor number, since insulin in association with FSH (Follicle stimulating Hormone) increases ovarian LH-binding capacity (Adashi et al., 1985; Davoren et al., 1986). In vitro studies have shown that insulin directly stimulates both mitosis and steroid production of cultured bovine granulosa (Gutierrez et al., 1999), theca (Stewart et al., 1995) and luteal cells (Mamluk et al., 1999). Willis et al. (1996) reported that insulin stimulates ovarian steroidogenesis by both granulosa and thecal cells, increasing production of androgens, estrogens and progesterone in vitro.

 

Selvaraju et al. (2002) recorded significantly higher plasma progesterone concentrations on day 10 of estrous cycle in insulin treated repeat breeding cattle than control. Sarath et al. (2008) reported a higher progesterone concentration in insulin-treated goats on days 12, 16, 20 and 24. Stewart et al. (1995) suggested that insulin enhances growth and proliferation of theca cells leading to production of progesterone. Increase in progesterone might be due to direct effect of insulin on corpus luteum or by increasing the ovulation rate. Donaldson and Hart (1981) reported that administration of estradiol-17-β increased blood insulin concentration in farm animals. Gong et al. (1994) and Monniaux and Pisselet (1992) reported that IGF-I enhances FSH stimulated estrogen and progesterone production in granulosa cells of bovines and ovines. Ramoun et al. (2007) found that pretreatment with insulin for 3 days before gonadotrophin-releasing hormone agonist injection increased the size of the largest follicle and the oestrus induction rate in buffaloes suffering from summer acyclicity.

 

Insulin stimulates the follicle production and estrogen synthesis (Gupta et al., 2010). Information regarding the effect of insulin administration during follicular and luteal phase of the cycle on conception rate in cows is limited. Therefore, the present study was designed to observe the effect of insulin on circulating LH and estradiol 17-β in crossbred cows.

 

MATERIALS AND METHODS

 

The present study was conducted on 48 crossbred cyclic cows aged 4-12 years, maintained at Instructional Dairy Farm, GBPUA&T, Pantnagar (Uttarakhand). Ethical approval was taken to conduct the study. Cows were divided into four groups (n=12), as per the day of insulin (0.25 IU/kg subcutaneously) treatment. It was day 0-3, 4-7 and 8-11 of estrous cycle in group I, II and III. Fourth group served as control and PBS was injected in equal amount to insulin suspension. Blood samples were collected on day 0, 5, 10, 16, and 21 of estrous cycle. Animals were detected in estrus and artificially inseminated with frozen semen.

 

Estradiol 17-β was assayed by RIA kits (Immunotech, France) as per guidelines of manufacturer of RIA kits. Luteinizing hormone (LH) level in blood was determined by ELISA. Samples stored at -20 oC were thawed and used for qualitative and quantitative determination of various hormones in blood.

 

The data obtained in the present study was analyzed statistically as per the methods described by Snedecor and Cochran (1994) and results were presented as Mean±SE.

 

RESULTS AND DISCUSSION

 

Serum estradiol 17-β concentration (pg/ml) in animals of different groups was estimated (Table 1). Analysis of variance indicated that E2 concentration on day 0, 5, 10, 16 and 21 in all groups and among group I, II, III and IV varied non-significantly on different days.

 

Serum estradiol 17-β concentration was also studied in insulin treated pregnant animals (Table 2). Blood level of E2 on day 5 varied significantly (P<0.01) among all the groups. E2 level in blood on day 5 was significantly (P<0.01 and P<0.05) lower in group I compared to Control and group III, respectively. For the same day it was significantly (P<0.05) lower in group II compared to group IV. E2 level in pregnant animals on day 5 was significantly lower in group I and II compared to IV and in group I compared to group III might be due to suppressing effect of insulin on immature follicles during luteal phase of estrous cycle. E2 concentration on day 0, 5, 10, 16 and 21 was studied and analysed between pregnant and non-pregnant animals of each group.

 

Table 1: Blood concentration (Mean±SE) of estradiol 17-β (pg/ml) during estrous cycle in insulin treated crossbred cows (n=12).

 

Treatment Groups	Days of estrous cycle
	Day 0	Day 5	Day 10	Day 16	Day 21
Group I	10.35± 3.15	7.82± 1.99	8.46± 1.79	8.35± 1.23	7.11± 1.73
Group II	12.53± 2.59	11.60± 3.28	11.65± 2.58	13.22± 3.29	11.49± 2.45
Group III	17.43± 3.05	13.94± 1.81	14.93± 2.49	16.13± 2.0	15.11± 3.26
Group IV	9.63± 1.72	12.89± 2.45	10.46± 2.22	12.72± 1.57	12.17± 2.38

 

Table 2: Blood concentration (Mean±SE) of estradiol 17-β (pg/ml) during estrous cycle in insulin treated pregnant crossbred cows

 

Days of estrous cycle	Treatment Groups
	Group I	Group II	Group III	Control Group
	Pregnant (n=7)	Pregnant (n=3)	Pregnant (n=5)	Pregnant (n=2)
Day 0	10.17± 5.54	9.19±4.34	22.67±6.45	18.94±1.55
Day 5	4.66± 2.05a	12.90± 4.99ac	15.44±3.46b	26.84±4.09b
Day 10	7.16±1.42	4.57±2.19	11.95±2.82	13.42±9.43
Day 16	8.27±1.53	11.51±6.97	17.92±2.40	17.48±0.27
Day 21	6.36±2.75	11.83±4.60	19.47±5.06	17.03±3.55

 

Means bearing different superscripts within the rows (a,b,c) differs significantly (P<0.05)

 

E2 might be in correlation with LH level as suggested by earlier researchers that the effect of insulin and LH on bovine granulosa cells is likely physiologically relevant (Spicer, 1998). Average concentrations of insulin and LH in beef and dairy cattle are usually less than 10 ng/ml (Richards et al., 1989; 1991) except at the time of the ovulatory surge of LH during which LH concentrations can achieve > 30 ng/ml (Richards et al., 1991). Studies in vivo have shown that insulin injections increase estradiol concentrations in follicular fluid of cattle (Simpson et al., 1994), and that estradiol concentrations in follicular fluid decrease after the LH surge in cattle (Voss and Fortune, 1993). Thus, insulin and LH may be physiologically relevant regulators of ovarian follicular estradiol production in cattle (Spicer, 1998).

 

Serum LH concentration (mIU/ml) in insulin treated crossbred cows during estrous cycle was estimated (Table 3) While it was also studied and analysed in insulin treated pregnant and non-pregnant animals (Table 4).

 

Table 3: Blood concentration (Mean±SE) of Luteinizing Hormone (m IU/ml) during estrous cycle in insulin treated crossbred cows

 

Treatment Groups	Days of estrous cycle
	Day 0	Day 5	Day 10	Day 16	Day 21
Group I (n=12)	3.41± 0.71	3.25± 0.62	2.04± 0.30	3.62± 0.96	3.18± 0.63
Group II (n=12)	3.33± 0.54	3.11± 0.89	3.74± 1.37	3.67± 0.91	3.53± 0.74
Group III (n=12)	4.08± 0.77	4.23± 0.97	4.07± 0.77	3.95± 0.94	4.58± 1.88
Group IV (control) (n=12)	3.21± 0.99	3.52± 0.70	5.20± 1.37	3.94± 0.66	6.07± 1.83

 

Table 4: Blood concentration (Mean±SE) of Luteinizing Hormone (m IU/ml) during estrous cycle in insulin treated pregnant crossbred cows

 

Days of estrous cycle	Treatment Groups
	Group I	Group II		Group III	Control Group
	Pregnant (n=7)		Pregnant (n=3)	Pregnant (n=5)		Pregnant (n=2)
Day 0	2.65±0.52		3.58±1.25	4.16±0.50ac		0.15±0.15
Day 5	2.97±0.71		2.33±0.51	4.46±0.68a		1.20±1.20
Day 10	2.30±0.43		3.47±1.62	4.56±1.56a		8.75±5.45
Day 16	4.46±1.52		3.14±0.87	1.76±0.81cb		5.30±2.39
Day 21	4.19±0.89		2.89±0.15	0.12±0.12b		1.45±1.45

 

Means bearing different superscripts within the column (a,b,c) differs significantly (P<0.05)

 

High level of LH in group I, II and III compared to control on day 0 and 21 was due to high rise of LH just before ovulation under the influence of E2 hence affecting corpus luteum formation as well as maturation and had positive effect on rise of progesterone too as in pregnant animals of control group the P4 level was also low since Luteinizing hormone (LH) is the major luteotropin in domestic ruminants (Niswender et al., 1985) and cattle (Oshea, 1987).

 

In pregnant animals the serum concentration of LH in group III on day 0, 5 and 10 was significantly (P<0.01) higher from day 21. Serum LH concentration on day 5, 10 was also significantly (P<0.05) higher from day 16. LH level decreases with advancement of pregnancy due to the negative feedback effect of P4 on LH secretion (Convey et al., 1977).

 

Serum concentration of LH on day 0, 5, 10, 16 and was analysed between pregnant and nonpregnant animals for all the groups. Its concentration on day 16 and 21 varied significantly (P<0.05) for group III. Concentration of LH on day 16 and day 21 in group III was significantly (P<0.05) lower in pregnant animals compared to nonpregnant animals. It was due to the negative feedback effect of P4 on GnRH and LH secretion (Hacket and Hafs 1969). Randel and Erb (1971) reported that plasma LH levels were decreased in those animals having increased plasma progesterone.

 

Level of LH in blood on day 10 was also significantly (P<0.05) higher in pregnant animals of group I compared to pregnant animals of control. LH supports the P4 secretion since luteinizing hormone treatment increased progesterone secretion in 6th–10th and 11th –16 th days of the estrous cycle as suggested by Rekawiecki (2007).

 

Conclusion

 

Insulin treatment declined the estrogen at day 5 and enhanced LH secretion up to day 10 reflecting its beneficial effect in corpus luteum survival and further conception.

 

Acknowledgments

 

Authors are thankful to the Dean, College of Veterinary and Animal Sciences and Joint Director, IDF, DES GBPUA & T, Pantnagar, for facilities and financial assistance extended during the course of study.

 

conflict of interest

 

There is no conflict of interest.

 

Authors contribution

 

All authors contributed equally.

 

References

 

• Adashi EY, Resnick CE, D’Ercole AJ, Svoboda ME, VanWyk JJ (1985). Insulin-like growth factors as intraovarian regulators of granulosa cell growth and function. Endocrine Rev. 6: 400-420. https://doi.org/10.1210/edrv-6-3-400

• Baithalu RK, Singh SK, Gupta C, Raja AK, Saxena A, Agarwal SK (2013). Insulin stimulates progesterone secretion to a greater extent than LH in early pregnant buffalo luteal cells cultured in vitro. Anim. Reprod. Sci. 142: 131-136. https://doi.org/10.1016/j.anireprosci.2013.09.004

• Convey EM, Beck TW, Neitzel RR, Bostwick EF, Hafs HD (1977). Negative feedback control of bovine serum luteinizing Hormone (lh) concentration from completion of the Preovulatory LH surge until resumption of luteal function. J. Anim. Sci. 46(4): 792-796. https://doi.org/10.2527/jas1977.454792x

• Davoren JB, Hsueh JW (1986). Growth hormone increases ovarian levels of immunoreactive somatomedinC/insulin-like growth factor I in vitro. Endocrinol. 118: 888-890. https://doi.org/10.1210/endo-118-2-888

• Donaldson IA, Hart IC (1981). Growth hormone, insulin, prolactin and total thyroxin in the plasma of sheep implanted with the anabolic steroid trenbole acetate alone or with oestradiol. Res. Vet. Sci. 30: 7-13.

• Garzo VG, Darrington JH (1984). Aromatase activity in human granulosa cells during follicular development and the modulation by follicle-stimulating hormone and insulin. Am. J. Obst. Gynaecol. 148: 657-62 https://doi.org/10.1016/0002-9378(84)90769-5

• Gong JG, Bramley TA, Wilmut I, Webb R (1993b). Effect of recombinant bovine somatotrophin on the superovulatory response to pregnant mare serum gonadotrophin in heifers. Biol. Reprod. 48: 1141-49. https://doi.org/10.1095/biolreprod48.5.1141

• Gong JG, McBride D, Bramley TA, Webb R (1994). Effect of recombinant bovine somatotrophin, insulin like growth factor-I and insulin on bovine granulose cell steroidogenesis in vitro. J. Reprod. Fertil. 110: 611-22.

• Gupta V, Thakur MS, Agrawal RG, Quadri MA, Shukla SN (2010). Effect of pre-treatment with Insulin on ovarian and fertility response in true anestrus buffaloes to Gonadotrophin–Releasing hormone. Buffalo Bull. 29(3): 172 – 179.

• Gutierrez CG, Gong JG, Bramley TA, Webb R (1999). Effects of genetic selection for milk yield on metabolic hormones and follicular development in postpartum dairy cattle. J. Reprod. Fertil. Abstract Series 24 Abstract 32.

• Hackett AJ, HD Hafs (1969). Pituitary and hypothalamic endocrine changes during the bovine estrous cycle. J. Anita. Sci. 28:531. https://doi.org/10.2527/jas1969.284531x

• Mamluk R, Greber Y, Meidan R (1999). Hormonal regulation of messenger ribonucleic acid expression for steroidogenic factor-1, steroidogenic acute regulatory protein, and cytochrome P450 side-chain cleavage in bovine luteal cells. Biol. Reprod. 60: 628–634. https://doi.org/10.1095/biolreprod60.3.628

• Monniaux D, Pisselet C (1992). Control of proliferation and differentiation of ovine granulose cells by insulin like growth factor-I and follicle stimulating hormone in vitro. Biol. Reprod. 46: 109-19. https://doi.org/10.1095/biolreprod46.1.109

• Niswender GD, Schwall RH, Fitz TA, Farin CE, Sawyer HR (1985). Regulation of luteal function in domestic ruminants: New concepts. Rec. Prog. Horm. Res. 41: 101-151.

• Oshea JD (1987). Heterogeneous cell types in the corpus luteum of sheep, goats and cattle. J. Reprod. Fert. 34: 71-85.

• Poretsky L, Piper B (1994). Insulin resistance, hyper-secretion of LH, and a dual-defect hypothesis for the pathogenesis of polycystic ovary syndrome. Obst. Gynaecol. 84: 613-21.

• Purkayastha RD, Shukla SN, Shrivastava OP, Kumar PR (2015). A comparative therapeutic management of anoestrus in buffaloes using insulin and GnRH. Vet. World. 2015 Jun. 8(6):804-7. https://doi.org/10.14202/vetworld.2015.804-807

• Randel RD, Erb RE (1971). Reproductive steroids in the bovine. VI. Changes and interrelations from 0 to 260 days of pregnancy. J. Anim. Sci. 33:115. https://doi.org/10.2527/jas1971.331115x

• Ramoun AA, Osman KT, Darwish SA, Karen AM, Gamal MH (2007). Effect of pretreatment with insulin on the response of buffaloes with inactive ovaries to gonadotropin-releasing hormone agonist treatment in summer. Reprod. Fertil. Develop. 19: 351-355 https://doi.org/10.1071/RD05097

• Rekawiecki R, Kotwica J (2007). Molecular regulation of progesterone synthesis in the bovine corpus luteum. Vet. Med. 52 (9): 405–412. https://doi.org/10.17221/1996-VETMED

• Richards MW, Wetterman RP, Sehoenmann H (1989). Nutritional Anestrus in beef cows : concentration of glucose and nonesterified fatty acids in plasma and insulin in serum. J. Anim. Sci. 67: 2354-2362. https://doi.org/10.2527/jas1989.6792354x

• Richards MW, Wetteman RP, Spicer LJ, Morgan GL (1991). Nutritional anestrus in beef cows: effects of body condition and ovariectomy on serum luteinizing hormone and insulin-like growth factor-I. Biol. Reprod. 44 : 961–966. https://doi.org/10.1095/biolreprod44.6.961

• Sarath T (2005). Follicular Dynamics, Endocrine and Nitric Oxide Profiles In Cyclic And Insulin Administered Acyclic Goats. M.V.Sc Thesis, IVRI, Izatnagar (U.P). morphological characteristics. Biol. Reprod. 56: 608–616.

• Selvaraju S, Agarwal SK, Karche SD, Srivastava SK, Majumdar AC, Shanker U (2002). Fertility responses and hormonal profiles in repeat breeding cows treated with insulin. Anim. Reprod. Sci. 73: 141-149. https://doi.org/10.1016/S0378-4320(02)00133-1

• Simpson RB, Chase JCC, Spicer LJ, Vernon RK, Hammond AC, Rae DO (1994). Effect of exogenous insulin on plasma and follicular insulin-like growth factor-I, insulin-like growth factor binding protein activity, follicular oestradiol and progesterone, follicular growth in superovulated Angus and Brahman cows. J. Reprod. Fertil. 102: 483-92. https://doi.org/10.1530/jrf.0.1020483

• Singh J Lalthazuali, SPS Ghuman, AK Pandey, GS Dhaliwal (2010). Impact of insulin treatment during post-insemination mid-luteal phase on luteal profile and conception rate in buffaloes. Indian J. Anim. Sci. 80 (9) (2010), Pp. 854-856.

• Snedecor GW, Cochran WG (1994). Statistical Methods, 8th Edn. Ames, Iowa State University Press. 503p.

• Spicer LJ, Alpizar A, Echternkamp SE (1993). Effects of insulin, insulin-like growth factor-I and gonadotrophins on bovine granulosa cell proliferation, progesterone production, oestradiol production and (or) insulin-like growth factor-I production in vitro. J. Anim. Sci. 71: 1232-41.

• Spicer LJ (1998). Tumor necrosis factor-_ (TNF_) inhibits steroidogenesis of bovine ovarian granulosa and thecal cells in vitro: involvement of TNF_ receptors. Endocrine 8: 109–115. https://doi.org/10.1385/ENDO:8:2:109

• Stewart RE, Spicer LJ, Hamilton TD, Keefer BE (1995. Effects of insulin-like growth factor I and insulin on proliferation and on basal and luteinizing hormone-induced steroidogenesis of bovine thecal cells: involvement of glucose and receptors for insulin-like growth factor I and luteinizing hormone. J. Anim. Sci. 73: 3719–3731. https://doi.org/10.2527/1995.73123719x

• Voss AK, Fortune JE (1993). Levels of messenger ribonucleic acid for cytochrome P450 17 alpha-hydroxylase and P450 aromatase in preovulatory bovine follicles decrease after the luteinizing hormone surge. Endocrinol. 132: 2239-45 . https://doi.org/10.1210/endo.132.5.8477668

• Willis D, Mason H, Gilling-Smith C, Franks S (1996). Modulation by insulin of FSH and luteinizing hormone actions in human granulosa cells of normal and polycystic ovaries. J. Clin. Endocrinol. Metabol. 81: 302-09. https://doi.org/10.1210/jc.81.1.302