Ovarian and endocrine characteristics during an estrous cycle in Angus, Brahman, and Senepol

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Ovarian and endocrine characteristics during an estrous cycle in Angus, Brahman,
and Senepol cows in a subtropical environment
P. Alvarez, L. J. Spicer, C. C. Chase, Jr, M. E. Payton, T. D. Hamilton, R. E. Stewart, A. C.
Hammond, T. A. Olson and R. P. Wettemann
J Anim Sci 2000. 78:1291-1302.
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Ovarian and endocrine characteristics during an estrous cycle in Angus,
Brahman, and Senepol cows in a subtropical environment1,2,3
P. Alvarez*, L. J. Spicer*, C. C. Chase, Jr.†,4, M. E. Payton‡, T. D. Hamilton*,
R. E. Stewart*, A. C. Hammond†,5, T. A. Olson§, and R. P. Wettemann*
Departments of *Animal Science and ‡Statistics, Oklahoma State University, Stillwater 74078; †Agricultural
Research Service, USDA, Brooksville, FL 34601-4672; and §Animal Science Department,
University of Florida, Gainesville 32610
ABSTRACT: Todeterminebreeddifferencesinovar- secondarysurgeofFSH(betweend1and2;d0=estrus)
ianfunctionandendocrinesecretion,dailyrectalultra- was greater in Angus than Brahman or Senepol cows
sonography was conducted on multiparous lactating (breed×day,P<.05).Betweend2and14oftheestrous
Angus(temperateBostaurus;n=12),Brahman(tropi- cycle, concentrations of progesterone, LH, IGF-II, and
calBosindicus;n=12),andSenepol(tropicalBostau- bindingactivitiesofIGFBP-3,IGFBP-2,andthe27-to
rus; n = 12) cows during an estrous cycle in summer. 29-kDaIGFBPinplasmadidnotdiffer(P>.10)among
Bloodwascollecteddailytoquantifyplasmaconcentra- breeds.ConcentrationsofGH,IGF-I,insulin,andPUN
tionsofFSH,LH,progesterone,estradiol,GH,insulin- weregreater(P<.001)andbindingactivitiesofthe22-
likegrowthfactor(IGF)-I,IGF-II,IGFbindingproteins kDa and 20-kDa IGFBP tended (P < .10) to be greater
(IGFBP), insulin, glucose, and plasma urea nitrogen inplasmaofBrahmanthaninAngusorSenepolcows.
(PUN). Numbers of small (2 to 5 mm), medium (6 to 8 Plasmaglucoseconcentrationsweregreater(P<.05)in
mm), and large follicles (≥ 9 mm) were greater (P < SenepolthaninBrahmanorAnguscows.Inconclusion,
.05) in Brahman than in Angus and(or) Senepol cows. Brahman(Bosindicus)andSenepolcows(tropicalBos
Length of the estrous cycle (SEM = .6 d) was similar taurus)hadgreaternumbersoffolliclesinallsizecate-
(P > .10) among Senepol (20.4 d), Angus (19.5 d), and goriesandgreaterdiameterofCLthanAngus(temper-
Brahman(19.7d)cows.Senepolcowshadgreater(P< ateBostaurus)cows.Theseovariandifferencesmaybe
.05)diametersofthecorpusluteum(CL)andadelayed duetochangesinthepatternofsecretionofFSH,insu-
regressionoftheCLascomparedwithAnguscows.The lin,IGF-I,andGHbutnotLH,IGF-II,orIGFBP-2or-3.
Key Words: Beef Cows, Binding Proteins, Cattle Breeds, Follicles, FSH, Insulin-Like Growth Factor
2000 American Society of Animal Science. All rights reserved. J. Anim. Sci. 2000. 78:1291–1302
Introduction 1984). In addition, puberty occurs at an older age in
Bos indicus than in Bos taurus cattle (Plasse et al.,
Differences in the reproductive characteristics be- 1968; Baker et al., 1989), and twinning rate is less in
tweenBostaurusandBosindicuscattleincludelonger BosindicusthaninBostauruscattle(Rutledge,1975).
gestationlengthforBrahman thanAnguscows(Foote Disparities in several reproductive characteristics, in-
etal.,1960;Reynoldsetal.,1980),andshorterandless- cluding gonadotropin (Rhodes et al., 1978) and steroid
intense estrus in Bos indicus than Bos taurus females (AdeyemoandHeath,1980;Segersonetal.,1984)secre-
(RhodesandRandel,1978;Galinaetal.,1982;Randel, tion, follicular growth and luteal development (Seger-
sonetal.,1984;Simpsonetal.,1994),and(or)thesecre-
1ThismanuscriptispublishedasFloridaAgric.Exp.Sta.Journal
SeriesNo.R-06661andapprovedforpublicationbythedirectorof 3Namesarenecessarytoreportfactuallyonavailabledata;how-
theOklahomaAgric.Exp.Sta.Thisresearchwassupportedinpart ever,theUSDAneitherguaranteesnorwarrantsthestandardofthe
underprojectH-2088(toL.J.Spicer). product, andthe useof thename byUSDA impliesno approvalof
2Appreciation is expressed to E. Achmadi, E. L. Adams, C. S. theproductortheexclusionofothersthatmayalsobesuitable.
Chamberlain,B.E.Keefer,E.V.Rooks,M.L.Rooks,andtheSTARS 4Correspondence: 22271 Chinsegut Hill Road. (phone: 352-796-
staff for technical assistance; the National Hormone and Pituitary 3385;fax:[phone];E-Mail:[email protected]fl.edu).
Program (Rockville, MD) for the IGF-I antiserum; Monsanto Com- 5Present address: USDA-ARS-PWA, 800 Buchanan St., Albany,
pany (St. Louis, MO) for the recombinant bovine IGF-II; and the CA94170.
OSURecombinantDNA/Protein ResourceFaciltyforthe useofits ReceivedJune30,1999.
molecularimagerandscanningdensitometer. AcceptedNovember12,1999.
1291
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1292 Alvarezetal.
tion of metabolites and metabolic hormones (Simpson taglandin-F (PGF; Lutalyse, Upjohn, Kalamazoo,
2α
etal.,1994;1997)mayaccountforthesedifferencesin MI). The estrous synchronization procedure consisted
reproductive performance. of an initial injection of 25 mg of PGF (i.m.), followed
Senepol cattle (tropically adapted Bos taurus) were byinjectionsof12.5mgofPGF11and12dlater(Santos
developed in the early 1900s on St. Croix, U.S. Virgin et al., 1988). Cows were visually observed for signs of
Islands, from a cross between West African (N’Dama) estrus for 1 h at 12-h intervals from 1 to 4 d after the
and European (Red Poll) breeds of cattle (Williams et lastinjectionofPGF.Sterilebullsfittedwithchinball
al., 1988). Several traits, such as heat tolerance and marker devices were used to assist in the detection of
diseaseandparasiteresistance,makeSenepolcattlea estrus.Onsetofestruswasconsideredtohaveoccurred
suitable breed for beef production in subtropical envi- when a cow stood to be mounted by another cow or
ronments (Hupp, 1981; Hammond and Olson, 1994; markerbull.Observationsforestrusresumed17dafter
Hammondetal.,1996).However,reproductivecharac- estrustodeterminethelengthoftheestrouscycle.Five
teristicsofSenepolfemaleshavenotbeenwellcharac- cows were removed from the original sample group of
terized.Ovarianfunctionandendocrinesecretionhave 36;onecowwasremovedduetothepresenceofacystic
notbeencomparedamongAngus,Brahman,andSene- follicle, two of the cows failed to exhibit behavioral es-
pol cows. Therefore, we set out to test the hypothesis trus, and two other cows had irregular estrous cycle
that ovarian function (follicular growth and luteal de- lengthsandirregularprogesteroneprofiles.Allexperi-
velopment)andendocrinesecretionofSenepolcowswas mental procedures were approved by the STARS Ani-
more like Brahman than Angus cows maintained in a mal Care and Use Committee.
subtropical environment. OvarianUltrasonography.Afterthesynchronizedes-
trouscycle,beginning ond0(estrus), ovarieswereex-
Materials and Methods amined by rectal ultrasonography (Aloka 210 ultra-
soundscannerequippedwitha7.5-MHzprobe,Corome-
Animals, Management, and Estrous Synchronization. tricsMedicalSystems,Inc.,Wallingford,CT)oncedaily
This study was conducted during summer (July and (beginning at 0800) during the second estrous cycle
August) of 1995 at the Subtropical Agricultural Re- followingthesynchronizedestrus.Imagesofeachovary
searchStation(STARS)locatednearBrooksville,Flor- were recorded using a VCR (General Electric
ida.Theaveragemaximumandminimumdailytemper- 1CVD5025X, General Electric Co., Portsmouth, VA).
atures during July were 32.6 and 22.8°C and during Tapes were played back, images were projected on a
Augustwere32.0and23.6°C,respectively;thehighest monitor, and a diagramdepicting the relative location
and lowest temperatures during July were 35.0 and of follicles ≥ 2 mm and the corpus luteum (CL) was
21.1°CandduringAugustwere35.5and21.1°C,respec- drawn for each ovary. Numbers and sizes of ovarian
tively; and total rainfall in July was 177 mm and in follicles and size of the CL (height, width, and cavity)
August was 360 mm. Total rainfall for August of 1995 were determined for each day. Ovulation was deter-
was132mmabovenormal.Twelvemultiparouslactat- mined by the disappearance of the dominant follicle
ingcowswererandomlyselectedfromeachoftheAngus andsubsequentformationofaCLatthesamelocation
(temperate Bos taurus; age = 6.5 ± .40 yr; BW = 417 ± in the ovary. Maximum diameter and growth rate of
9.4 kg; body condition score = 5.2 ± .21, 1 = thin to 9 = the dominant follicle of the first follicular wave and of
obese; dayspostpartum =148 ±5.0 d),American Gray theovulatoryfolliclewerealsoestimated.Growthrate
Brahman (tropical Bos indicus; age = 6.9 ± .43 yr; BW of the dominant follicle was determined from the day
= 490 ± 15.7 kg; body condition score = 5.2 ± .27; days thedominantfolliclewasfirstidentifiedtothedaythat
postpartum = 134 ± 3.0 d) and Senepol (tropical Bos the diameter of the follicle no longer increased more
taurus; age = 6.3 ± .22 yr; BW = 464 ± 11.0 kg; body than 1 mm. Growth rate of the ovulatory follicle was
conditionscore=4.4±.23;dayspostpartum=120±5.1 determined from first detection to ovulation. Number
d) cow herds at STARS. Within breed, no more than of follicular waves during the estrous cycle was deter-
two cows with the same sire were selected and 100% mined using the identity method (Knopf et al., 1989),
of the Angus, 67% of the Brahman, and 100% of the in which individual follicles on each ovary (> 5 mm)
Senepol cows also had calved the previous year. Cows were identified and assigned a position relative to the
andcalvesweremaintainedinadrylot(withaccessto otherovarianstructurespresent(i.e.,CLandfollicles).
shade) as one group and given ad libitum access to Awaveoffolliculargrowthwasidentifiedbytheforma-
mixed rhizoma perennial peanut (Arachis glabrata tionofadominantfollicleandagroupofgrowingfolli-
Benth)-bahiagrass (Paspalum notatum) hay (large cles (subordinates) associated with the dominant folli-
roundbales;10.3%CPand47.8%TDN,DMbasis)and cle. The day of emergence was defined as the last day
custommineralmixture(25to32%salt,15to18%Ca, agroupoffollicleswere4mmasindicatedbyincreasing
5to8%P, ≥.94%Fe,≤.15%Fl,≥.10% Cu,≥.01%Co, diameters on subsequent days. The dominant follicle
and .0010 to .0015% Se). During the study, ADG was was defined as the largest follicle present on either
−.3 ± .08 kg/d for Angus, −.2 ± .08 kg/d for Brahman, ovarybefored12,andasubordinatefolliclewasdefined
and .1 ± .12 kg/d for Senepol. Calves were allowed to as onethat originated fromthe same follicularpool as
suckleadlibitum.Estruswassynchronizedusingpros- the dominant follicle and was indicated by: 1) its first
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Folliculardynamicsamongbreedsofbeefcows 1293
detection within 2 d of the first detection of the domi- assays were 9.9% and 13.5%, respectively. Sensitivity
nantfollicle;and2)hadasubsequentincreaseindiame- of the assay was .12 ± .01 ng/mL.
ter.Numbersoffolliclesweredividedintothreedistinct Concentrationofestradiolinplasmawasdetermined
sizeclassesforanalyses:small(2to5mm),medium(6 in daily samples using a solid-phase Serono Estradiol
to 8 mm), and large (≥ 9 mm) follicles. MAIAassaykit(PolymedcoInc.,CortlandtManor,NY)
Blood Collection and Metabolite and Hormone Analy- after extraction with ethyl acetate as previously de-
ses.Duringthesynchronizedestrouscycleandthesub- scribed (Vizcarra et al., 1997). Intra- and interassay
sequentnaturalestrouscycle(whenovarianultrasound CV for 9 assays were 15.0% and 24.3%, respectively.
was conducted), a single blood sample was collected Sensitivity of the assay was .63 ± .06 pg/mL.
dailyfromeachcow(beginningat0800)byjugularveni-
Growthhormoneconcentrationinplasmawasquan-
puncture into 9-mL blood collection tubes containing
tified in daily samples by a double-antibody RIA as
EDTA (Monovette Sarstedt, Newton, NC). Blood sam-
previously described (Yelich et al., 1995). Intra- and
ples were placed on ice immediately after collection,
interassay CV for five assays were 8.0% and 15.8%.
andplasmawasseparatedbycentrifugation(4°C,1,750
Sensitivity of the assay was 8.4 ± 1.4 ng/mL.
×gfor30min).Plasmasampleswerefrozenandstored
PlasmaconcentrationofLHwasdeterminedindaily
at −20°C until metabolite and hormone analyses were
samples by a double-antibody RIA as previously de-
performed.Foreachmetaboliteandhormoneanalysis,
scribed (Bishop and Wettemann, 1993). Intra- and in-
all breeds were represented in each assay.
terassay CV for five assays were 12.1% and 19.2%, re-
Plasma concentration of glucose was determined in
spectively.Sensitivityoftheassaywas.31±.09ng/mL.
daily samples by an automated colorimetric method
Ligand Blots. Plasma IGF binding proteins (IGFBP)
(TechniconAutoAnalyzerII;IndustrialMethod339-19,
were analyzed in samples collected on d 10 by one di-
Technicon Industrial Systems, Tarrytown, NY) based
mensionalSDS-PAGE,aspreviouslydescribed(Echter-
ontheglucoseoxidaseprocedureasdescribedbyGoch-
nkampetal.,1994;Simpsonetal.,1997).Briefly,sam-
man and Schmitz (1972). Concentration of PUN was
determined in daily samples by an automated colori- ples (4.0 (cid:1)L added to 21.0 (cid:1)L of buffer) were heat-
metric procedure (Technicon AutoAnalyzer II; Indus- denaturedandthenseparatedona12%polyacrylamide
trial Method 339-01, Technicon Industrial Systems, gel via electrophoresis. After separation, proteins in
Tarrytown, NY) based on the diacetyl monoxime gels were electrophoretically transferred to nitrocellu-
method described by Marsh et al. (1965). lose, and ligand-blotted overnight with [125I]IGF-II.
Concentration of IGF-I in plasma collected every Afterwashingandthenexposuretox-rayfilmat−70°C
otherdaywasdeterminedbydouble-antibodyRIAafter for48h,bandintensityonautoradiographswasdeter-
acid-ethanol extraction (16 h at 4°C) as described pre- minedusingaPDIModelDNA35scannerandQuantity
viously (Echternkamp et al., 1990). Intra- and in- One (Version 2.4) software for quantification by scan-
terassay CV for five plasma IGF-I assays were 12.1% ning densitometry.
and 23.4%, respectively. Sensitivity of the assay, de- StatisticalAnalyses.Theexperimentwasacompletely
fined as 90% of total binding, was 3.7 ± .6 ng/mL. randomized design with repeated measures. The data
To compare follicular phase and luteal phase IGF- wereanalyzedusingPROCMIXED(Littelletal.,1996),
II levels, plasma concentration of IGF-II for samples with sources of variation including breed, cow within
collected on d 0 and d 10 was determined by a double- breed (error term for breed), day, breed × day interac-
antibody RIA after formic acid-acetone extraction as
tion,andresidual.Anautoregressivewithlagequalto
describedpreviously(Stewartetal.,1996).Allsamples
one model was used to model the covariance structure
wereevaluatedinoneassay;theintrassayCVwas2.7% oftherepeatedmeasurements.Ifthebreed×dayinter-
and sensitivity was 50 ng/mL.
actionwassignificant,simpleeffectsofbreedwereana-
Plasma concentration of progesterone was deter-
lyzedusingtheSLICEoptionfortheLSMEANSstate-
mined in daily samples using a solid-phase RIA kit
ment.Satterthwaite’sapproximationwasusedforcal-
(Diagnostics Products Corporation, Los Angeles, CA)
culation of the degrees of freedom of the pooled error
as previously described (Stewart et al., 1996). Intra-
term.Ifthebreed×dayinteractionwasnotsignificant,
andinterassayCVfor11assayswere4.4%and10.6%,
respectively. Sensitivity of the assay was .05 ± .002 the main effects, if significant, were analyzed using
LSMEANS with the PDIFF option (SAS, 1989). For
ng/mL.
certainvariables(estradiol,CLdiameter,progesterone,
Concentration of FSH in plasma was quantified in
LH), the maximum value over the estrous cycle was
daily samples by a double-antibody RIA as previously
described(Vizcarraetal.,1997).Intra-andinterassay also analyzed. The resulting design was a completely
CV for 10 assays were 11% and 27.6%, respectively. randomized design and PROC GLM was used to ana-
Sensitivity of the assay was .05 ± .01 ng/mL. lyze these data with LSMEANS reported (SAS, 1989).
Plasma concentration of insulin was measured in Numberofwavesofovarianfolliculardevelopmentwas
dailysamplesusingasolid-phaseRIAkit(ICNPharma- analyzed by PROC FREQ using chi-square analysis
ceuticalsInc.,CostaMesa,CA)asdescribedpreviously (SAS,1989).Dataarepresentedasleastsquaresmeans
(Simpsonetal.,1994).Intra-andinterassayCVfor11 ± SEM.
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1294 Alvarezetal.
(Figure1B);therewasneitheradayeffectnorbreed×
dayinteraction(P>.10).Thenumberoflarge(≥9mm)
follicles was greater (P < .05) in Brahman (1.6 ± .2)
thaninAngus(.9±.2)orSenepol(1.2±.2)cows(Figure
1C),andtherewasnobreed×dayinteraction(P>.10).
There was a day effect (P < .001) on number of large
follicles.Numberoflargefolliclesinallbreedsincreased
(P < .05) from d 1 (day of ovulation) to d 20 of the
estrous cycle.
Maximum diameter of the dominant follicle of the
first wave of follicular development was less (P < .01)
inAnguscowsthaninBrahmanorSenepolcows(Table
1). Growth rate of the first dominant follicle tended to
be greater (P < .10) in Brahman cows than in Angus
andSenepolcows(Table1).Maximumdiameterofthe
ovulatoryfolliclewasgreater(P<.01)inBrahmanthan
in Angus or Senepol cows (Table 1). No differences (P
>.10)ingrowthrateoftheovulatoryfolliclewerefound
among Angus, Brahman, and Senepol cows (Table 1).
AlthoughAngusandBrahmancowshadpredominantly
twowavesoffolliculardevelopmentduringtheestrous
cycle(72.7%and55.6%,respectively)andSenepolcows
hadmostly(70%)threewavesoffolliculardevelopment
during the estrous cycle (Table 2), these breed differ-
ences were not significant (P > .10). Length of the es-
trouscycledidnotdiffer(P>.10)amongSenepol(20.4
± .6 d), Angus (19.5 ± .6 d), and Brahman (19.7 ± .6 d)
cows. Among breeds, length of the estrous cycle was
longer(P<.005)incowswiththreewavesoffollicular
development(20.9±.47d)thanincowswithtwowaves
of follicular development (18.7 ± .44 d) during the es-
trous cycle.
Luteal Development. There was a breed × day (P <
.06)interactionforCLdiameteracrossd0to20.Regres-
sion of the CL occurred earlier in the estrous cycle of
AngusandBrahmancowsthaninSenepolcows(Figure
2A). If CL diameter was analyzed for only d 2 to 14 of
the estrous cycle, the breed × day interaction was not
significant (P > .10). The average diameter of the CL
between d 2 to 14 was greater (P < .001) in Brahman
(16.4 ± .3 mm) and Senepol (16.3 ± .3 mm) than in
Figure 1. Number of small (2 to 5 mm; Panel A), me- Angus (15.0 ± .3 mm) cows. The diameter of the CL
dium (6 to 8 mm; Panel B), and large (≥ 9 mm; Panel C) increased (P< .001)from d2 to 8in allbreeds (Figure
follicles in Angus, Brahman, and Senepol cows during 2A). Maximum diameter of the CL was greater (P <
anestrouscycleasdeterminedbydailyrectalultrasonog- .05)inSenepol(22.2±.3mm)thaninAngus(19.6±.2
raphy. Standard errors averaged over days were 3.9, .7, mm)cowsandtended(P<.10)tobegreaterinBrahman
and.2forsmall,medium,andlargefollicles,respectively. (21.3±.3mm)thaninAnguscows.MaximumCLdiam-
eterwassimilar(P>.10)betweenBrahmanandSene-
pol cows.
Results Endocrine Profiles.Therewasabreed×day(P<.07)
interactionforplasmaconcentrationofestradiolacross
Follicular Dynamics and Function. Number of small d0to20oftheestrouscycle.Thepreovulatoryincrease
(2 to 5 mm) follicles was greater (P < .05) in Brahman of estradiol was earlier in the estrous cycle of Angus
(39±4)thaninAngus(21±4)orSenepol(33±4)cows andBrahmancowsthanSenepolcows(Figure3).When
and was greater (P < .001) in Senepol than in Angus plasmaconcentrationofestradiolwasanalyzedford2
cows (Figure 1A); there was neither a day effect nor to 14 of the estrous cycle to assess “basal” estradiol
breed × day interaction (P > .10). Number of medium secretion, the breed× day interaction stillexisted (P <
(6 to 8 mm) follicles was less (P < .05) in Angus (2.3 ± .06); this was due to an increase in plasma estradiol
.7)thaninBrahman(5.0±.7)orSenepol(3.9±.7)cows observed between d 2 and 5 in Angus and Brahman
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Folliculardynamicsamongbreedsofbeefcows 1295
Table 1. Characteristics of dominant and ovulatory follicles during an estrous cycle in
Angus, Brahman, and Senepol cows as determined by rectal ultrasonography
Firstdominantfollicle Ovulatoryfollicle
Maximum Growthrate, Maximum Growthrate,
Breed diameter,mm mm/d diameter,mm mm/d
Angus 11.4 ± .6a 1.18 ± .12c 12.8 ± .4a 1.39 ± .10
Brahman 15.3 ± .6b 1.55 ± .12d 15.6 ± .5b 1.35 ± .11
Senepol 13.9 ± .5b 1.22 ± .12c 13.6 ± .4a 1.46 ± .10
a,bWithinacolumn,meanslackingacommonsuperscriptletterdiffer(P<.01).
c,dWithinacolumn,meanslackingacommonsuperscriptletterdiffer(P<.10).
cows but not in Senepol cows (Figure 3). Maximum × day interaction and mean plasma concentration of
concentrationofestradiolinplasmadid notdiffer(P> progesterone did not differ among breeds. The concen-
.10) among Angus (9.1 ± 1.4 pg/mL), Brahman (8.9 ± tration of progesterone increased (P < .001) in plasma
1.6 pg/mL), and Senepol (8.7 ± 1.4 pg/mL) cows. fromd0to12inallthreebreeds(Figure2B).Maximum
There was a breed × day (P < .01) interaction for plasma concentration of progesterone did not differ (P
plasma concentrationof progesterone across d0 to 20. >.10)amongAngus(4.3±.33ng/mL),Brahman(4.4±
Luteal regression, indicated by a decrease in plasma .37 ng/mL), and Senepol (5.2 ± .33 ng/mL) cows.
progesterone, occurred earlier in Angus and Brahman There was a breed × day interaction (P < .05) for
cows than in Senepol cows (Figure 2B). When plasma plasma concentrations of FSH across d 0 to 20 of the
concentration of progesterone was analyzed for d 2 to estrouscycle.TheincreasesofFSH betweend1and2
14 of the estrous cycle, there was no significant breed of the estrous cycle were more pronounced (P < .005)
in Angus than in Brahman and Senepol cows (Figure
4A).MaximumplasmaFSHconcentrationachievedbe-
tweend0and4averaged1.18,.61,and.67±.10ng/mL
for Angus, Brahman, and Senepol cows, respectively.
Duringd0to20oftheestrouscycle,plasmaconcentra-
tion of FSH averaged .62, .45, and .44 ± .03 ng/mL for
Angus, Brahman, and Senepol cows, respectively.
There was a breed × day interaction (P < .01) for
concentrations of LH in plasma across d 0 to 20 of the
estrouscycle(Figure4B).Ifdataforplasmaconcentra-
tionofLHwereanalyzedford2to14oftheestrouscycle
toassess“basal”LHsecretion,therewerenosignificant
effects of breed, day, or breed × day. During d 2 to
14 of the estrous cycle, plasma concentration of LH
averaged4.3,4.2,and4.4±.5ng/mLforAngus,Brah-
Figure 2. Diameter of the corpus luteum (CL; Panel
A) as determined by daily rectal ultrasonography and Figure3.PlasmaconcentrationsofestradiolinAngus,
plasmaconcentrationofprogesterone(PanelB)inAngus, Brahman,andSenepolcowsasdeterminedbyRIA.Blood
Brahman,andSenepolcowsduringanestrouscycle.Stan- samples were collected daily. Standard errors averaged
darderrorsaveragedoverdayswere.9mmand.2ng/mL overdayswere.4,.4,and.3pg/mLforAngus,Brahman,
for CL diameter and plasma progesterone, respectively. and Senepol cows, respectively.
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1296 Alvarezetal.
Figure 4.Plasma concentrationsof FSH (PanelA) and
LH (Panel B) in Angus, Brahman, and Senepol cows as
determinedby RIA.Bloodsampleswere collecteddaily.
Figure 5. Plasma concentrations of growth hormone
Standard errors averaged over days were .2 ng/mL and
(GH; Panel A) and insulin-like growth factor I (IGF-I;
.5 ng/mL for FSH and LH, respectively.
PanelB)inAngus,Brahman,andSenepolcowsasdeter-
mined by RIA. Blood samples were collected daily for
GHandeveryotherdayforIGF-I.Standarderrorsaver-
man,andSenepolcows,respectively.Maximumplasma
aged over days were 6.9 ng/mL and 2.6 ng/mL for GH
LHconcentrationdidnotdiffer(P>.10)amongbreeds
and IGF-I, respectively.
and averaged 31.7, 30.7, and 22.1 ± 8.15 ng/mL for
Angus, Brahman, and Senepol cows, respectively.
ConcentrationofGHinplasmafromd0to20ofthe (P < .001) plasma concentrations of IGF-I than Angus
estrous cycle was greater (P < .001) in Brahman (42.2 cows. Neither day nor the breed × day interaction af-
±2.2ng/mL)thaninAngus(17.4±2.1ng/mL)orSene- fected plasma concentrations of IGF-I (P > .10).
pol (20.1 ± 2.0 ng/mL) cows (Figure 5A). There was no Plasma concentration of IGF-II determined on d 0
dayeffectorbreed×dayinteraction(P>.10)observed and10withineachbreeddidnotdiffer(P>.10)between
for plasma concentration of GH. d 0 (173 ± 7 ng/mL) and d 10 (169 ± 7 ng/mL). Plasma
Concentration of IGF-I in plasma samples collected concentration of IGF-II on d 0 and 10 did not differ (P
everyotherdayfromd0to20oftheestrouscyclewas >.10)amongbreedsandaveraged166,176,and171±
greater (P < .001) in Brahman (22.9 ± .9 ng/mL) than 9ng/mLforAngus,Brahman,andSenepolcows,respec-
in Senepol (14.5 ± .8 ng/mL) and Angus (10.0 ± .8 ng/ tively. There was no breed × day (P > .10) interaction
mL) cows (Figure 5B). Senepol cows also had greater for plasma concentration of [125I]IGF-II.
Table 2. Number of waves of ovarian follicular development and length of the estrous
cycle in Angus, Brahman, and Senepol cows as determined by rectal ultrasonography
2-Wavecycles 3-Wavecycles
Number
Breed ofcows n(%) Cyclelength,d n(%) Cyclelength,d
Angus 11 8(72.7) 18.3 ± .6 3(27.3) 20.7 ± .9
Brahman 9 5(55.6) 18.6 ± .7 4(44.4) 20.8 ± .8
Senepol 10 3(30.0) 19.3 ± .9 7(70.0) 21.4 ± .6
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Folliculardynamicsamongbreedsofbeefcows 1297
(Figure 7B); plasma glucose concentrations also dif-
fered (P < .05) between Brahman and Senepol cows.
There was a day effect (P < .01), but no breed × day
interactiononplasmaconcentrationofglucose,indicat-
ingthatthedecreaseinglucoseconcentrationbetween
d0and8oftheestrouscyclewassimilaramongbreeds
(Figure 7B).
Plasma urea nitrogen concentration from d 0 to 20
oftheestrouscyclewasgreater(P<.001)inBrahman
cows(15.4±.2mg/dL)thaninAngus(11.8±.2mg/dL)
orSenepol(14.6±.2mg/dL)cows,andwasalsogreater
Figure 6. Binding activity of IGFBP-2, IGFBP-3, a 27-
to29-kDaIGFBP,anda20-and22-kDaIGFBPinAngus,
Brahman,and Senepolcows ond10 ofthe estrouscycle
as determined by ligand blotting.
Ligand blotting with [125I]IGF-II revealed at least
fiveformsofIGFBP; 40-to44-kDa(IGFBP-3),34-kDa
(IGFBP-2),27-to29-kDa,22-kDa,and20-kDa.Binding
activitiesexpressedinarbitrarydensitometricunitsof
IGFBP-3,IGFBP-2,andthe27-to29-kDaIGFBP(Fig-
ure6A)didnotdiffer(P>.10)amongbreedsandaver-
aged 13.1 ± 1.0, 7.4 ± .5, and 6.5 ± .5, respectively, on
d 10 of the estrous cycle. Brahman cows tended (P <
.10)tohavegreater22-kDaand20-kDaIGFBPbinding
activity (3.5 ± .4 and 1.0 ± .2 unit) than Angus (2.6 ±
.4and.5±.2unit)orSenepol(2.3±.4and.3±.2unit)
cows (Figure 6B).
Plasmaconcentrationofinsulinfromd0to20ofthe
estrouscyclewasgreater(P< .001)inBrahman(.68±
.05ng/mL)thaninAngus(.44±.05ng/mL)andSenepol
(.49±.05ng/mL)cows,butdidnotdifferbetweenAngus
andSenepolcows(Figure7A).Therewasneitheraday Figure 7. Plasma concentrations of insulin (Panel A),
nor breed × day effect (P > .10) on plasma insulin. glucose(PanelB),andplasmaureanitrogen(PUN;Panel
Plasma Metabolites. Concentration of glucose in C)inAngus,Brahman,andSenepolcows.Bloodsamples
plasma from d 0 to 20 of the estrous cycle was greater werecollecteddaily.Standarderrorsaveragedoverdays
(P < .001) in Senepol (76.7 ± .6 mg/dL) and Brahman were .2 ng/mL, 3.1 mg/dL and .9 mg/dL for insulin,
(74.5±.7mg/dL)thaninAngus(68.8±.6mg/dL)cows glucose, and PUN, respectively.
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1298 Alvarezetal.
(P < .001) in Senepol than in Angus cows (Figure 7C). Fortune,1988;Gintheretal.,1989;Zeitounetal.,1996),
There was a day effect (P < .001) but no breed × day and artificially extending the luteal phase with exoge-
interaction (P > .10) on PUN concentration, indicating nous progesterone has resulted in estrous cycles with
that the decrease in PUN concentration between d 0 four or five waves of follicular growth (Sirois and For-
and 20 of the estrous cycle was similar among breeds tune, 1990).
(Figure 7C). In spite of the dramatic differences in the number
and growth patterns of follicles that existed among
Discussion breeds, no differences in maximum plasma concentra-
tionofestradiolwereobserved.However,thesignificant
Ovarian follicular growth as indicated by number breed×dayinteractionforplasmaconcentrationofes-
of follicles in all categories, growth rate of the first tradiol revealed that there was an increase in concen-
dominant follicle to develop during the estrous cycle, trationofestradiolond5oftheestrouscycleofAngus
andmaximumdiameterofthefirstdominantandovu- andBrahmancowsbutnotSenepolcows;thiswaslikely
latory follicles was greater in Brahman (Bos indicus) due to the development of the first dominant follicle.
thaninAngus(temperateBostaurus)cowsofthepres- There was an earlier preovulatory increase in concen-
ent study. Rather than being Brahman-like, follicular tration of estradiol in Angus and Brahman cows than
growthinSenepol(tropicalBostaurus)cowswasinter- Senepolcows;however,peak(maximum)concentration
mediate and between that observed for Brahman and of estradiol did not differ among breeds. In contrast,
Anguscows,andthuswerejectouroriginalhypothesis. Segerson et al. (1984) found that the concentration of
Theseresultsareconsistentwithpreviousstudiesindi- estradiolfromd7to17oftheestrouscyclewaslessin
cating that numbers of antral follicles are greater in BrahmanthaninAnguscows.Reasonsforthisinconsis-
Brahman than in Angus cows (Segerson et al., 1984; tency are unclear but may be due to 1) the different
Simpson et al., 1994). The hormonal stimulus for in- seasons that samples were collected between studies
creased follicular growth in Brahman vs Senepol vs (i.e.,summerinthepresentstudyvsfallintheSegerson
Angus cows is discussed in detail later but is most etal.(1984)study);2)Angusheifersbeinglesstolerant
closely associated with increased secretion of IGF-I. toheatthanBrahmanandSenepolheifers(Hammond
Toourknowledge,thisisthefirststudyinwhichthe etal.,1996);and(or)3)Segersonetal.,(1984)including
wave pattern of follicular growth in Senepol cows was pregnant and nonpregnant cows in their analysis. A
determined. Senepol cows had mostly three waves of greater concentration of estradiol in plasma of Brah-
follicular development during their estrous cycle, mancowsthaninAnguscowswouldbeexpecteddueto
whereasthemajorityofAngusandBrahmancowshad thegreaternumberoffolliclesintheformer.However,
twowaves. ThepercentageofBrahman cowswithtwo Simpsonetal.,(1994)reportedthatfolliclesfromBrah-
waves of follicular development (56%) was greater in mancowstreatedwithgonadotropinstosuperstimulate
thepresentstudythanthe38%reportedbyZeitounet folliculardevelopmentcontainlessestradiolthanthose
al.(1996),butlessthanthe84%reportedbyFigueiredo ofAnguscows;thiscouldexplainwhymaximumplasma
et al. (1997). Reasons for these discrepancies are un- concentrationofestradioldidnotdifferbetweenBrah-
clearbutmaybeduetothefactthatthepresentstudy manandAnguscowsinthepresentstudy.Whyplasma
was conducted during summer (July and August) in concentration of estradiol was similar between Brah-
Florida, whereas Zeitoun et al. (1996) conducted their man and Angus cows in spite of a greater number of
study during the spring (May) and Fall (October) in follicles in Brahman cows is uncertain. Perhaps the
Texas, and Figueiredo et al. (1997) conducted their two-fold greater concentration of IGF-I in plasma of
study during winter (July and August) in Brazil. Sea- BrahmanthanAnguscowsinthepresentstudyinhib-
sonaldifferencesinthenumberandsizeofantralfolli- ited insulin-stimulated estradiol production by granu-
cles of Brahman cows have been reported (Lammoglia losa cells of small and large follicles as previously re-
et al., 1996). Differences in nutritional status of the ported (Spicer et al., 1994). Thus, Brahman cows with
cowsbetweenstudiescouldalsoaccountforthediscrep- greaterconcentrationofIGF-Imayhavereducedestra-
ancies. In the present study, cows were allowed free diolproductionfromfolliclesandneedmorefolliclesto
access to rhizoma perennial peanut hay, whereas Zei- yield the same plasma concentration of estradiol as
toun et al. (1996) and Figueiredo et al. (1997) supple- Angus cows.
mented cows with concentrate. The delayed luteal re- In addition to greater concentration of IGF-I, the
gression of the Senepol cows may account for the pre- greater follicular activity in Brahman cows could be
dominance of three waves of follicular development explained by the greater mean concentration of GH
during estrous cycles in Senepol cows. Ginther et al. and(or)insulinthaninAnguscows.Thegreatersecre-
(1989)suggestedthatthenumberofwavesoffollicular tion ofGH and IGF-I inBrahman than inAngus cows
developmentduringtheestrouscycleincattleisregu- agrees with previous studies (Simpson et al., 1994;
lated by the length of the luteal phase. Generally, es- 1997). Unfortunately, the frequency of blood sampling
trouscycleswiththreeorfourwavesoffolliculardevel- usedinthepresentstudy(i.e.,daily)didnotlenditself
opment have longer luteal phases than estrous cycles to an analysis of GH pulse frequency or amplitude. In
with two waves of follicular development (Sirois and vitro,insulinandIGF-Iarepotentstimulatorsofbovine
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Folliculardynamicsamongbreedsofbeefcows 1299
granulosa and thecal cell proliferation, whereas GH, The greater luteal growth in Brahman and Senepol
LH,andFSHhavelittleornoeffect(Spiceretal.,1993; cowsvsAnguscouldbeexplainedbythegreaterconcen-
Stewart et al., 1995; Spicer and Stewart, 1996). How- tration of GH and(or) IGF-I. Injections of GH in cattle
ever,exogenousinsulinhadnoeffectonthenumberof that increased serum IGF-I concentration also in-
antral follicles in Angus and Brahman cows treated creased size of CL and concentration of progesterone
withgonadotropinstosuperstimulateovarianfollicular inplasma(Schemmetal.,1990;GalloandBlock,1991;
development(Simpsonetal.,1994).Inheifersandlac- Lucyetal.,1994).Invitro,IGF-Istimulatesprogester-
tating dairycows treatedwith bovine GH,an increase oneproductionbyculturedbovinelutealcells(Schams
inthenumberofsmallfollicleswascorrelatedwithan et al., 1988; Sauerwein et al., 1992; Chakravorty et
increase in serum GH and IGF-I (Gong et al., 1992; al.,1993),andreducedlutealprogesteronesecretionis
1993; Lucy et al., 1992). However, in spite of the in- associated with decreased plasma IGF-I in dairy cows
creasednumbersoffolliclesafterGHtreatment,others (Spiceretal.,1990).LutealtissueinBosindicus(Brah-
(Schemm et al., 1990; Gong et al., 1991) did not find man)andBostaurus(Hereford)cowshavesimilarhis-
anydifferencesbetweencontrolandGH-treatedcowsin tologicalandmorphologicalcharacteristics,suchasor-
systemicconcentrationofestradiol,whichisconsistent ganization, apparent population of cells per area, and
with findings of the present study. Because treatment celltypespresent(Irvinetal.,1978).Invitro,however,
with GH increases peripheral concentrations of GH, luteal tissue from Brahman cows secretes less proges-
IGF-I, and insulin (Gong et al., 1991; 1993; Stanko et terone than luteal tissue from Hereford cows (Rhodes
al., 1994), determining the hormonal cause for in- et al., 1982). The latter observation could explain why
creasedfolliculargrowthandsteroidogenesisafterGH inourstudyBrahmancowshadlargerCLthanAngus
treatment can be difficult. butplasmaconcentrationofprogesteronedidnotdiffer
To our knowledge this is the first time CL diameter between these breeds; perhaps Brahman cows needed
inBostaurusandBosindicuscowshavebeencompared largerCLtoproducethesameamountofprogesterone
using ultrasonography. We found that luteal growth becauseoftheirlargerbodymass.Inadditiontodirect
was greater in Brahman and Senepol than in Angus stimulatoryeffectsofIGF-IonCLgrowthandsteroido-
cows.However,smallerCLmaybepresentinBosindi- genesis, direct effect of GH on the CL seems likely,
custhaninBostaurusheifers(Irvinetal.,1978;Rhodes since bovine CL contained more GH receptor mRNA
etal.,1982;Segersonetal.,1984).Theseinconsistencies and GH receptor protein than other reproductive tis-
between the results of present and previous studies sues (Lucy et al., 1993; Kirby et al., 1996).
may be due to differences in timing and frequency of Ligand blotting with [125I]IGF-II revealed at least
luteal measurements during the estrous cycle, and(or) fiveformsofIGFBP: 40-to44-kDa(IGFBP-3),34-kDa
differences in season in which the studies were con- (IGFBP-2), 27- to 29-kDa, 22-kDa, and 20-kDa. Breed
ducted. differences in IGFBP-3 and -2 activity were not found
Inspiteof10%greaterCLdiameterinBrahmanand inthepresentstudy,andthisagreeswiththestudyof
Senepol than in Angus cows, there was no difference Simpson et al. (1997), in which no differences were
in concentration of progesterone among breeds before found in IGFBP-2 and total IGFBP binding activity
d14oftheestrouscycle.Thisfindingagreeswithprevi- betweenovariectomizedAngusandBrahmancows,but
ous studies in which no difference was detected in itisinconsistentwithSimpsonetal.(1994),whofound
plasma concentration of progesterone (Rhodes et al., 10%greatertotalplasmaIGFBPactivityinAngusthan
1982; Diaz et al., 1986), but it contrasts with others inBrahmancowstreatedwithgonadotropinstosuper-
studiesthatfoundlowerconcentrationofprogesterone stimulatefolliculardevelopment.Simpsonetal.(1997)
(AdeyemoandHeath,1980;Randel,1984;Segersonet found greater plasma IGFBP-3 levels in estradiol-
al.,1984)inBosindicusthanBostauruscattle.Differ- treatedovariectomizedBrahmanthanAnguscows.Dif-
encesinthesamplingfrequency,seasonandclimateat ferences in season, physiological state of animals, and
timeofsampling,andageandphysiologicalstate(e.g., frequencyofsamplingmayaccountfortheinconsistenc-
lactating or not) of the animals may account for these ies of results between studies. Growth hormone has
inconsistencies. Heat stress affects production of pro- been implicated in the regulation of IGFBP-2 and -3
gesterone,because,inHolsteincows,serumconcentra- (Stanko et al., 1994). However, despite the differences
tionsofprogesteronebetweend6and18oftheestrous incirculatingGH,nosignificantdifferencesinIGFBP-
cycle were less during summer than during spring 2 or-3 were found amongbreeds in thepresent study.
(Howelletal.,1994).Inaddition,lutealcontentofpro- Differencesinthehypothalamic-hypophysealaxisex-
gesterone was greater in Holstein cows exposed than istbetweenBosindicus andBostauruscattle(Rhodes
incowsnotexposedtoforcedventilation(Younasetal., et al., 1978). Ovariectomized Brahman cows produced
1993).Inagreementwithpreviousstudies(Kastelicet lessLHinresponsetoGnRHthanHerefordcows(Grif-
al.,1990;RajamahendranandTaylor,1990;Ribaduet finandRandel,1978).Also,Brahmanheifershaveless
al.,1994),CLdiameterandconcentrationofprogester- LHduringthepreovulatorysurgeofLH,anearlierLH
one was positively correlated in Angus (r = .70), Brah- surge, and they have a shorter interval from estrus to
man (r = .73), and Senepol (r = .75) cows between d 0 ovulationthanHerefordcows(Randel,1984).Wefound
and 14 of the estrous cycle of the present study. no difference in mean concentration of LH between d
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