C57BL/6J mouse superovulation: schedule and age optimization to increase oocyte yield and reduce animal use

Abstract Superovulation protocols have been described for different mouse strains, however the numbers of animals used are still high and still little information is known about hormone administration schedules and estrous cycle phases. In this study, we aimed to optimize a superovulation protocol by injecting 5 IU of pregnant mare serum gonadotropin followed by 5 IU of hCG 48 h later, using three different schedules related to the beginning of the dark cycle (3, 5 and 7 pm) in a light cycle of 7 am to 7 pm, with light on at 7 am. C57BL/6J mice at 3, 4 and 5 weeks of age were used and the estrous cycle phase for times of PMSG and hCG injections was also analyzed. Total oocyte number was counted in the morning after hCG injection. Hormones given at 3 weeks of age at 3 pm (59 ± 15 oocytes) and 7 pm (61 ± 10 oocytes) produced a significantly higher oocyte number compared with oocytes numbers collected from females at the same age at 5 pm (P = 0.0004 and <0.0001 respectively). Females at 4 and 5 weeks of age produced higher numbers of oocytes when superovulated at 7 pm. No statistical differences between females at different phases of the estrous cycle were found. These results showed that in C57BL/6J mice, hormones should be given at 3 or 7 pm for females at 3 weeks of age, however older females should be superovulated closer to the beginning of the dark cycle to reduce female mouse use and increase the numbers of oocytes produced per female.

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Transient circadian internal desynchronization after light-dark phase shift in monkeys

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1977.232.1.r31 ◽

1977 ◽

Vol 232 (1) ◽

pp. R31-R37 ◽

Cited By ~ 1

Author(s):

M. C. Moore-Ede ◽

D. A. Kass ◽

J. A. Herd

Keyword(s):

Body Temperature ◽

Saimiri Sciureus ◽

Cycle Phase ◽

Dark Phase ◽

Dark Cycle ◽

Water Excretion ◽

Urinary Potassium ◽

Timing System ◽

Internal Desynchronization ◽

Circadian Timing System

In four conscious chair-acclimatized squirrel monkeys (Saimiri sciureus) studied with lights on (600 lx) from 0800 to 2000 h daily (LD 12:12), prominent 24-h rhythms in feeding, drinking, activity, body temperature, and urinary potassium, sodium, and water excretion were seen. When the monkeys were subjected to 36 h of darkness followed by 36 h of light each variable demonstrated a circadian rhythm which was not passively dependent on the light-dark cycle. After the 24-h light-dark cycle was abruptly phase-delayed by 8 h, all the rhythms resynchronized with the new light-dark cycle phase, demonstrating that light-dark cycles are an effective zeitgeber. However, the resynchronization of the rhythms of feeding, drinking, activity, and body temperature was 90% complete within approximately 2 days while the 90% resynchronization of the urinary rhythms took approximately 5 days. These results suggest that the circadian timing system in S. sciureus may consist of several spontaneously oscillating units which can become transiently uncoupled during pertubations of environmental time cues.

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Stage of the estrous cycle at the time of pregnant mare's serum gonadotropin injection affects pre-implantation embryo development in vitro in the mouse

Molecular Reproduction and Development ◽

10.1002/mrd.10132 ◽

2002 ◽

Vol 62 (3) ◽

pp. 312-319 ◽

Cited By ~ 12

Author(s):

Juan J. Tarín ◽

Sonia Pérez-Albalá ◽

Vanessa Gómez-Piquer ◽

Carlos Hermenegildo ◽

Antonio Cano

Keyword(s):

Estrous Cycle ◽

Embryo Development ◽

Serum Gonadotropin ◽

Development In Vitro

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Reversed light–dark cycle and cage enrichment effects on ethanol-induced deficits in motor coordination assessed in inbred mouse strains with a compact battery of refined tests

Behavioural Brain Research ◽

10.1016/j.bbr.2011.05.030 ◽

2011 ◽

Vol 224 (2) ◽

pp. 259-271 ◽

Cited By ~ 14

Author(s):

Elizabeth Munn ◽

Mark Bunning ◽

Sofia Prada ◽

Martin Bohlen ◽

John C. Crabbe ◽

...

Keyword(s):

Motor Coordination ◽

Inbred Mouse ◽

Mouse Strains ◽

Inbred Mouse Strains ◽

Dark Cycle

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Effect of adrenalectomy and corticosterone replacement on meal patterns of Zucker rats

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1985.249.5.r584 ◽

1985 ◽

Vol 249 (5) ◽

pp. R584-R594 ◽

Cited By ~ 5

Author(s):

M. R. Freedman ◽

T. W. Castonguay ◽

J. S. Stern

Keyword(s):

Weight Gain ◽

Food Intake ◽

Body Weight Gain ◽

Zucker Rats ◽

Meal Frequency ◽

Light Cycle ◽

Dark Cycle ◽

Meal Patterns ◽

Total Food Intake ◽

Total Food

Male obese and lean Zucker rats were adrenalectomized (ADX) or sham-operated at 10 wk of age. Approximately 16 wk later, patterns of food intake were monitored by computer-interfaced top loading balances. Data were collected from ADX rats before, during, and after access to a corticosterone-supplemented saline solution (20 micrograms/ml). Although total food intake during the precorticosterone treatment period was not different between ADX and sham controls, ADX resulted in attenuation of light cycle food intake, primarily via decreased meal frequency. With steroid replacement, light cycle meal frequency and food intake increased. Despite comparable self-administered dose (20.33 +/- 0.89 vs. 17.05 +/- 1.2 mg corticosterone/period, obese vs. lean), obese ADX rats were more responsive to steroid than were lean ADX rats. This increased responsiveness was reflected by a 30% increase in food intake and 60% increase in body weight gain of obese ADX rats during replacement. Lean ADX rats exhibited no change in total food intake or weight gain with replacement. Further, during corticosterone treatment, obese ADX rats increased meal frequency, total food intake, and consumption of large meals (greater than or equal to 4 g) during the dark cycle. Significant postprandial correlations were found only in obese ADX rats, both with and without replacement during the dark cycle. These results suggest adrenal glucocorticoids have a minimal effect on food intake and meal patterns in lean Zucker rats but significantly alter intake and meal patterns in obese rats.

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TrkB-expressing neurons in the dorsomedial hypothalamus are necessary and sufficient to suppress homeostatic feeding

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1815744116 ◽

2019 ◽

Vol 116 (8) ◽

pp. 3256-3261 ◽

Cited By ~ 15

Author(s):

Guey-Ying Liao ◽

Clint E. Kinney ◽

Juan Ji An ◽

Baoji Xu

Keyword(s):

Leptin Receptor ◽

Critical Role ◽

Light Cycle ◽

Dark Cycle ◽

Neuronal Tracing ◽

Dorsomedial Hypothalamus ◽

Adult Mice ◽

Treatment Of Obesity ◽

Control Body ◽

Necessary And Sufficient

Genetic evidence indicates that brain-derived neurotrophic factor (BDNF) signaling through the TrkB receptor plays a critical role in the control of energy balance. Mutations in the BDNF or the TrkB-encoding NTRK2 gene have been found to cause severe obesity in humans and mice. However, it remains unknown which brain neurons express TrkB to control body weight. Here, we report that TrkB-expressing neurons in the dorsomedial hypothalamus (DMH) regulate food intake. We found that the DMH contains both glutamatergic and GABAergic TrkB-expressing neurons, some of which also express the leptin receptor (LepR). As revealed by Fos immunohistochemistry, a significant number of TrkB-expressing DMH (DMHTrkB) neurons were activated upon either overnight fasting or after refeeding. Chemogenetic activation of DMHTrkB neurons strongly suppressed feeding in the dark cycle when mice are physiologically hungry, whereas chemogenetic inhibition of DMHTrkB neurons greatly promoted feeding in the light cycle when mice are physiologically satiated, without affecting feeding in the dark cycle. Neuronal tracing revealed that DMHTrkB neurons do not innervate neurons expressing agouti-related protein in the arcuate nucleus, indicating that DMHTrkB neurons are distinct from previously identified LepR-expressing GABAergic DMH neurons that suppress feeding. Furthermore, selective Ntrk2 deletion in the DMH of adult mice led to hyperphagia, reduced energy expenditure, and obesity. Thus, our data show that DMHTrkB neurons are a population of neurons that are necessary and sufficient to suppress appetite and maintain physiological satiation. Pharmacological activation of these neurons could be a therapeutic intervention for the treatment of obesity.

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Determination of the estrous cycle phases of rats: some helpful considerations

Brazilian Journal of Biology ◽

10.1590/s1519-69842002000400008 ◽

2002 ◽

Vol 62 (4a) ◽

pp. 609-614 ◽

Cited By ~ 712

Author(s):

F. K. MARCONDES ◽

F. J. BIANCHI ◽

A. P. TANNO

Keyword(s):

Estrous Cycle ◽

Cell Types ◽

Short Length ◽

Female Rats ◽

Cycle Phase ◽

Objective Lens ◽

Vaginal Smear ◽

Vaginal Secretion ◽

Cellular Types

The short length of the estrous cycle of rats makes them ideal for investigation of changes occurring during the reproductive cycle. The estrous cycle lasts four days and is characterized as: proestrus, estrus, metestrus and diestrus, which may be determined according to the cell types observed in the vaginal smear. Since the collection of vaginal secretion and the use of stained material generally takes some time, the aim of the present work was to provide researchers with some helpful considerations about the determination of the rat estrous cycle phases in a fast and practical way. Vaginal secretion of thirty female rats was collected every morning during a month and unstained native material was observed using the microscope without the aid of the condenser lens. Using the 10 x objective lens, it was easier to analyze the proportion among the three cellular types, which are present in the vaginal smear. Using the 40 x objective lens, it is easier to recognize each one of these cellular types. The collection of vaginal lavage from the animals, the observation of the material, in the microscope, and the determination of the estrous cycle phase of all the thirty female rats took 15-20 minutes.

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Pharmacological evidence for β2-adrenoceptor in right atria from stressed female rats

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y99-031 ◽

1999 ◽

Vol 77 (6) ◽

pp. 432-440 ◽

Cited By ~ 13

Author(s):

R C Spadari-Bratfisch ◽

I N Santos ◽

LCM Vanderlei ◽

F K Marcondes

Keyword(s):

Estrous Cycle ◽

Model Equation ◽

Female Rats ◽

Cycle Phase ◽

Response Curves ◽

Chronotropic Response ◽

Site Model ◽

Stressed Female ◽

Right Atria ◽

Shock Stress

The purpose of the present study was to demonstrate a physiological response to TA2005, a potent β2-adrenoceptor (β2-AR) selective agonist, in right atria isolated from stressed female rats under the influence of the estrous cycle. We obtained concentration-response curves to the agonist in the presence and in the absence of selective antagonists in right atria isolated from female rats submitted to three daily foot-shock sessions (30 min duration, 120 pulses of 1.0 mA, 1.0 s, applied at random intervals of 5-25 s) and sacrificed at estrus or diestrus. Our results showed that the pD2 values of TA2005 were not influenced by estrous cycle phase or foot-shock stress. However, in right atria from stressed rats sacrificed during diestrus, the concentration-response curve to TA2005 was biphasic, with a response being obtained at concentrations of 0.1 nM, whereas during estrus no response was observed at doses lower than 3 nM. ICI118,551, a β2-AR antagonist, abolished the response to nanomolar concentrations of TA2005 in right atria from stressed rats at diestrus, with no changes in agonist pD2 values in right atria from control rats (7.47 ± 0.09, p > 0.05) but a 3-fold decrease in pD2 values of TA2005 in right atria from foot shock stressed rats (7.90 ± 0.07, p [Formula: see text] 0.05). Concentration-response curves to TA2005 in the presence of ICI118,551 were best fitted by a one-site model equation. The β1-AR antagonist, CGP20712A, shifted to the right only the second part of the concentration-response curves to the agonist, unmasking the putative β2-AR-mediated response to the agonist in tissues isolated from stressed rats at diestrus. Under this condition, concentration-response curves to the agonist were best fitted by a two-site model equation. pD2 and maximum response of TA2005 interaction with β1- and putative β2-adrenoceptor components were calculated. Schild analyses gave a pKB value for CGP20712A that was typical for the interaction with β1-AR in each experimental group. pKB values for ICI118,551 could not be obtained in stressed rats sacrificed at diestrus since Schild plot slopes were lower than 1.0. In right atria from control rats, ICI118,551 pKB values were similar to reported values for the interaction of the antagonist with β1-AR. These results confirm that a heterogenous β-AR population mediating the chronotropic response to catecholamines can be demonstrated in right atria from foot shock stressed female rats sacrificed at diestrus. The stress-induced response seems to be mediated by the β2-AR subtype. Right atria from rats sacrificed during estrus are protected against stress-induced alterations on the homogeneity of β-AR population.Key words: foot-shock stress, TA2005, ICI118,551, CGP20712A, estrus, diestrus.

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Effect of estrous cycle phase on clinical pathology values in beagle dogs

Veterinary Clinical Pathology ◽

10.1111/j.1939-165x.2011.00392.x ◽

2012 ◽

pp. n/a-n/a ◽

Cited By ~ 1

Author(s):

Cynthia J. Willson ◽

Sundeep A. Chandra ◽

Carie L. Kimbrough ◽

Holly L. Jordan

Keyword(s):

Estrous Cycle ◽

Clinical Pathology ◽

Cycle Phase ◽

Beagle Dogs

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Sex differences in the circadian control of hamster wheel-running activity

AJP Regulatory Integrative and Comparative Physiology ◽

10.1152/ajpregu.1983.244.1.r93 ◽

1983 ◽

Vol 244 (1) ◽

pp. R93-R105 ◽

Cited By ~ 10

Author(s):

F. C. Davis ◽

J. M. Darrow ◽

M. Menaker

Keyword(s):

Sex Differences ◽

Sex Difference ◽

Wheel Running ◽

Phase Shifts ◽

Light Cycle ◽

Dark Cycle ◽

Running Activity ◽

Wheel Running Activity ◽

Activity Onset ◽

Males And Females

The circadian pacemaker that underlies the wheel-running activity of hamsters was studied in males and females. Sex differences were found in the mechanism by which the pacemaker entrains to light-dark cycles and in the timing of activity onset. When exposed to a light-dark cycle with a period of 24.75 h (with 1 h of light/cycle), males show a greater ability to maintain entrainment than do females. This difference in the upper limit of entrainment appears due to a sex difference in the magnitude of light-induced phase shifts. A small difference in free-running period may also contribute to the sex difference in entrainment. Two weeks after castration of adults, the sex difference in entrainment is not affected, indicating that the difference does not depend on circulating gonadal steroids or on estrous cyclicity of the female. However, castration of females at an early age increases their ability to entrain, whereas long-term castration of males seems to reduce entrainment ability. During entrainment to a 24-h light-dark cycle (LD 14:10), females were found to begin their daily activity before males and before castrated females. This difference is consistent with a sex difference in the magnitude of light-induced phase shifts and in entrainment of the pacemaker. However, evidence is given that the sex difference in activity onset might also be caused by a sex difference in the relationship of locomotor activity to the pacemaker in intact males and females.

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