guide histologi vaginal smear

http://tpx.sagepub.com/Toxicologic Pathology

http://tpx.sagepub.com/content/36/3/375The online version of this article can be found at:

DOI: 10.1177/0192623308315665

2008 36: 375 originally published online 25 April 2008Toxicol PatholF. Russell Westwood

The Female Rat Reproductive Cycle: A Practical Histological Guide to Staging

Published by:

http://www.sagepublications.com

On behalf of:

Society of Toxicologic Pathology

can be found at:Toxicologic PathologyAdditional services and information for

http://tpx.sagepub.com/cgi/alertsEmail Alerts:

http://tpx.sagepub.com/subscriptionsSubscriptions:

http://www.sagepub.com/journalsReprints.navReprints:

http://www.sagepub.com/journalsPermissions.navPermissions:

What is This?

- Apr 25, 2008Proof

- Jun 3, 2008Version of Record >>

by guest on November 5, 2011tpx.sagepub.comDownloaded from

http://tpx.sagepub.com/

http://tpx.sagepub.com/content/36/3/375

http://www.sagepublications.com

http://www.toxpath.org/

http://tpx.sagepub.com/cgi/alerts

http://tpx.sagepub.com/subscriptions

http://www.sagepub.com/journalsReprints.nav

http://www.sagepub.com/journalsPermissions.nav

http://tpx.sagepub.com/content/36/3/375.full.pdf

http://tpx.sagepub.com/content/early/2008/04/25/0192623308315665.full.pdf

http://online.sagepub.com/site/sphelp/vorhelp.xhtml


375

The Female Rat Reproductive Cycle: A Practical Histological Guide to Staging

F. RUSSELL WESTWOOD

Global Safety Assessment, AstraZeneca, Macclesfield, Cheshire SK10 4TG, United Kingdom

ABSTRACT

During preclinical investigations into the safety of drugs and chemicals, many are found to interfere with reproductive function in the female rat.This interference is commonly expressed as a change in normal morphology of the reproductive tract or a disturbance in the duration of particularphases of the estrous cycle. Such alterations can be recognized only if the pathologist has knowledge of the continuously changing histologicalappearance of the various components of the reproductive tract during the cycle and can accurately and consistently ascribe an individual tract to aparticular phase of the cycle. Unfortunately, although comprehensive reports illustrating the normal appearance of the tract during the rat estrous cyclehave been available over many years, they are generally somewhat ambiguous about distinct criteria for defining the end of one stage and the begin-ning of another. This detail is absolutely essential to achieve a consistent approach to staging the cycle. For the toxicologic pathologist, this reportillustrates a pragmatic and practical approach to staging the estrous cycle in the rat based on personal experience and a review of the literature fromthe last century.

Keywords: rat; reproductive; cycle; estrous; proestrus; estrus; metestrus; diestrus; histological; staging.

beginning of another. This detail is absolutely essential toachieve a consistent approach to staging the cycle. Indeed, someauthors provide only a single description of regions of the tractfrom rats with a four-day estrous cycle, collected at a specifictime on each day, as typical of a particular phase (Yuan andFoley 2002), which can lead to inconsistency when assessingprogressive and cyclical changes. My aim here is to illustrate,for the toxicologic pathologist, a pragmatic and practicalapproach to staging the estrous cycle in the rat based on per-sonal experience and literature references from the last century.The images presented (hematoxylin-and-eosin–stained sectionsprocessed by standard histological techniques) are examplesfrom young adult undosed Wistar rats (Alderley Park strain) butare entirely consistent with, and illustrative of, other strains.I have also included descriptions of the appearance of the tractduring aging. Some details of the associated hormonal changesare included in relation to the morphological changes, particu-larly for the aging rat, but they are necessarily cursory, as this isnot the main purpose of the report. For a complete review of thehormonal correlates, see The Physiology of Reproduction, editedby Knobil and Neil (1994).

THE ESTROUS CYCLE

Estrus was used first by Heape (1900) as a Latin adaptationof the Greek word oistros, meaning gadfly, sting, or frenzy todescribe the “special period of sexual desire of the female”

INTRODUCTION

During preclinical investigations into the safety of drugs andchemicals, many are found to interfere with the reproductivefunction of the female rat (Yuan and Foley 2002). This interfer-ence is commonly expressed as a change in normal morphologyof the reproductive tract or a disturbance in the duration ofparticular phases of the estrous cycle. These alterations can berecognized only if the pathologist has knowledge of the contin-uously changing histological appearance of the various com-ponents of the reproductive tract during the cycle and canaccurately and consistently ascribe an individual tract to a par-ticular phase of the cycle. Indeed, functional alterations may beapparent only following such an assessment and may manifestthemselves by a change in the number of animals in a particu-lar phase of the cycle, a disturbance in the coordinated mor-phology of the components of the tract, or abnormal appearanceof a particular region. Unfortunately, although comprehensivereports illustrating the normal appearance of the tract duringthe rat estrous cycle have been available over many years(Hubscher et al. 2005; Long and Evans 1922; Marshall 1960;Yuan and Foley 2002), they are generally somewhat ambiguousabout distinct criteria for defining the end of one stage and the

Address correspondence to: Dr. F. Russell Westwood, AstraZeneca,Mereside, Alderley Park, Macclesfield, Cheshire SK10 4TG, United Kingdom;e-mail: [email protected].

Toxicologic Pathology, 36: 375-384, 2008Copyright © 2008 by Society of Toxicologic PathologyISSN: 0192-6233 print / 1533-1601 onlineDOI: 10.1177/0192623308315665

Invited Reviews



(Freeman 1994). He also used: anestrus, nonbreeding seasonwhen reproductive organs are quiescent; proestrus, animal com-ing on heat; metestrus, in the absence of conception, when estruschanges in the reproductive tract subside; and diestrus, repro-ductive tract prepares for receipt of the ovum. Clearly, these“periods,” although they are accompanied by morphologicalchanges, they are not described by them, although the morpho-logical appearances of the reproductive organs of the rat havebeen well characterized under this general behavioral scheme, asnoted in the introduction. Vaginal smears are also widely used forthis purpose; their characteristics have been described in detailelsewhere (Hubscher et al. 2005; Long and Evans 1922; Maedaet al. 2000). However, they will also be discussed alongside themorphological features of the estrous stages under “Staging theCycle” below in support of the proposed method.

The onset of puberty in the female rat results from a cascadeof events following establishment of a pulsatile luteinizinghormone (LH) release after the fourth postnatal week (approxi-mately thirty days of age) that leads to ovarian maturation(Andrews and Ojeda 1981). Before this time, the reproductivetract is inactive. This change in LH release is apparent eight tonine days before the first proestrus, and this period of change inmode of LH release is considered anestrus (Urbanski and Ojeda1985). The first proestrus, estrus, and diestrus periods then follow(Advis et al. 1979; Ojeda et al. 1976).

Ovulation occurs in the young adult laboratory rat every fourto five days throughout the year (Ojeda and Urbanski 1994).The normal length of the estrous cycle has been investigated overmany decades. Long and Evans (1922) used approximately 2000young adult rats derived from descendents of a cross betweenseveral white females and a wild grey caught in Berkeley,California, and fed on table scraps and other supplements,including raw liver. They found marked individual variations,with cycle lengths from three to thirty-eight days, but, excludingthose of more than eight days, the average was 4.8 days, and thisduration is reasonably consistent with findings from subsequentinvestigators (Astwood 1939; Blandau et al. 1941). Based onvaginal smears, the duration of the individual components ofthe estrous cycle for rats with a four- or five-day cycle areproestrus, twelve to fourteen hours; estrus, twenty-five totwenty-seven hours; metestrus, six to eight hours; and diestrus,fifty-five to fifty-seven hours (Astwood, 1939; Hartman 1944;Long and Evans 1922; Mandl 1951). However, as noted above,many authors refer to the day of the cycle, with each periodhaving its own day, and those in a five-day cycle generallyshowing either an extra day of vaginal cornification (extra dayof estrus) or an extra day of leukocyte infiltration (extra day ofdiestrus) (vom Sall et al. 1994).

STAGING THE CYCLE

Complete longitudinal sections of the vagina and cervix,transverse sections of the mid portion of both uterine horns, andmedial sections of both ovaries are the minimal requirement foran adequate evaluation.

Observation of the coordinated morphology of the vaginaand uterus is key to consistent staging of the estrous cycle, andthis will be the main focus of the descriptions provided here.

The stage of the reproductive cycle cannot easily be determinedfrom the appearance of the ovarian follicles in a species like therat, which has a short estrous cycle. Follicles in virtually allstages of development are generally present within the ovariesat all phases of the cycle. However, the formation, progression,and regression of the corpora lutea are somewhat synchronized,and they can be used as an aid (Yuan and Foley 2002). It is ofnote that three or more generations of corpora lutea may bepresent in an individual ovary from the preceding ovulatorycycles, as each corpora lutea persists morphologically fortwelve to fourteen days (Long and Evans 1922; Yuan and Foley2002). Thus, the use of corpora luteal morphology as an accuratetool for estrous staging can be confusing. But the presence ofparticular morphologies of corpora lutea can be used to supportstaging, even though this is not definitive.

Table 1 summarizes the characteristics of the vaginal, uterine,and ovarian morphology during the four estrous stages seen inthe normally cycling adult rat. These cycles are described below.

Diestrus

Figure 1 illustrates the diestrus stage.

Vagina: At the start of diestrus, the vaginal epithelium is at its low-est level of approximately three to seven cells thick consistingsimply of the stratum germinativum. The stratum germinativumconsists of stratum basale as a single layer of columnar epithe-lial cells and an outer stratum spinosum as multiple layers ofpolyhedral cells. There is a variable infiltration by leukocytes.A reduction in the infiltration of leukocytes and a notableepithelial cell proliferation occurs toward the end of the phasewith thickening of the epithelium, but with no clear stratumgranulosum. The formation of a stratum granulosum is the defin-ing characteristic for the practical staging of the end of diestrusand the beginning of proestrus. The vaginal smear is consistentwith this histological appearance, which is characterized bylittle mucus with some leukocytes, nucleated basophilic cells,and occasional vacuolated cells.

Uterus: At the start of diestrus, the uterus is small and inactive,and the horns lack a prominent vasculature and generally showa slit-like lumen. They are lined by a low cuboidal or columnarepithelium showing occasional degenerate cells. There are fewmitoses at the start of the stage and the glands are particularlyinactive, but there is some increase in activity during the pro-gression of the phase. Also, toward the end of the phase, a slightedema of the stroma adjacent to the endometrial epithelium canbe seen.

Ovaries: By diestrus, in the ovary, the newly formed corporalutea from the previous ovulation have attained their maximalsize, and this is the best ovarian marker for diestrus, althoughdegenerate corpora lutea will also be present. Vacuoles arecommonly present, particularly in the cells in the center of theselarge corpora lutea, indicative of active steroidogenesis, and earlyfibrous tissue formation may be seen in what was previouslythe central, fluid-filled cavity.

376 WESTWOOD TOXICOLOGIC PATHOLOGY



Proestrus

Proestrus is illustrated in Figure 2.

Vagina: Formation of the stratum granulosum over the stratumgerminativum of the vaginal epithelium, consisting of flattenedepithelial cells containing many keratohyalin granules, marksthe start of proestrus. The vaginal epithelium shows mitoticfigures throughout the stage, although they are less numerousat the end. Following the early formation of the stratum granu-losum, there is a progressive development of the superficialmucoid layer (stratum mucification according to Yuan and Foley[2002], or rete mucosum according to Hubscher et al. [2005]),characterized by layers of cuboidal to ovoid cells with mucin-containing cytoplasmic vacuoles, and the formation of a stratumcorneum of dense, cornified cells. There is little if any degen-eration or desquamation during the early- or mid-proestrusperiod, and occasional infiltrates of leukocytes are seen. At theend of the stage, the epithelium is fully cornified and generallyshows a superficial mucoid layer exhibiting some desquama-tion of mucoid cells. The vaginal smear is again consistent withthis histological appearance, showing a disappearance of leuko-cytes and the presence of sheets of, or isolated, nucleated epithe-lial cells, which become progressively acidophilic with theappearance of cornified cells.

Uterus: During this stage, the uterine endometrial liningprogresses to large cells, forming a tall cuboidal to columnarepithelium. There are frequent mitoses with only limited or noepithelial cell degeneration of the glands and lining epithelium

(the appearance of notable endometrial epithelial cell degen-eration marks the end of proestrus/start of estrus) and littleinflammatory cell infiltration, although mitotic figures disappearat the end of the stage. The endometrial vasculature becomesmore prominent, and the stroma can show some edema, withthe lumen generally becoming markedly dilated toward theend of the stage.

Ovaries: Ovarian corpora lutea are often degenerate, with centralfibrous tissue formation, and the cells commonly contain cyto-plasmic vacuoles.

Estrus

Figure 3 illustrates estrus.

Vagina: In the vagina, there is a loss of mitotic figures and aprogressive shedding of the superficial mucoid and cornifiedlayers during estrus, with a reduction in the height of the epithe-lium, and cell debris is present in the lumen. There is also a vari-able and progressive leukocyte infiltration. The end of the stageis characterized by detachment of the cornified epithelium,although some may persist, particularly adjacent to the vaginalopening. So, if there is a predominance of attached cornifiedepithelium, then the stage is still “estrus.” Virtually completedetachment of the cornified epithelium of the vagina marks theend of estrus and the start of metestrus. The vaginal smear showsnon-nucleated cornified cells, which by late estrus have dimin-ished when leukocytes appear. Also, large basophilic epithelialcells are present in the smear.

Vol. 36, No. 3, 2008 STAGING THE FEMALE RAT REPRODUCTIVE CYCLE 377

TABLE 1.—Summary of the defining histological features of the rat female reproductive tract during diestrus, proestrus, estrus, and metestrus.

Vagina Uterus Ovaries

Diestrus Start defined by epithelium at lowest level with Small, avascular, slit-like lumen. Lined by low Large corpora lutea. May be finely variable leukocyte infiltration. Subsequent columnar epithelium. Initially few mitoses, but vacuolated. Fibrous tissue formation epithelial proliferation and thickening (no some increase during phase. Only occasional in central cavity.clear stratum granulosum) with reduction in degenerate cells. Stromal edema at end of stage.leukocyte infiltration.

Proestrus Mitotic figures present. Occasional polymorphs. Epithelium cuboidal to columnar. Mitoses present Corpora lutea often degenerate. Little if any degeneration or desquamation. in epithelial cells with little or no degeneration Cytoplasmic vacuoles generally Formation of stratum granulosum (defines and little inflammatory cell infiltration. present. Fibrous tissue proliferation start), superficial mucoid layer and stratum Dilatation, particularly toward end of stage. in central cavity.corneum progressively. At end of stage, fully cornified and generally showing superficial mucoid layer with some desquamation of mucoid cells.

Estrus Progressive shedding of superficial mucoid and Start of estrus defined by appearance of Degenerate corpora lutea often present. cornified layers. Reduction in height of notable degeneration/necrosis of epithelial Some small corpora lutea with epithelium. Cell debris present. Loss of mitotic cells, glands generally first. Loss of mitotic basophilic cell cytoplasm, central figures. Progressive leukocyte infiltration. activity. Leukocyte infiltration. Dilatation may fluid-filled cavity, and no fibrous

persist to late estrus. tissue.Metestrus Start defined by virtually complete detachment Continued degeneration of endometrial epithelial Corpora lutea may still contain fluid

of cornified layer (generally residual squames cells. Return of mitotic activity; both (mitotic cavity. Smaller than at diestrus. in lumen). Continued desquamation with loss of activity and degeneration) seen together. Slightly basophilic cells. Generally stratum granulosum and upper germinativum. devoid of fibrous tissue.Leukocyte infiltration.



Uterus: Changes in the endometrial epithelium define the startof estrus, with the appearance of cellular degeneration/necrosisin the glands first, followed by the lining epithelium, whichbecomes quite marked. It is accompanied by a loss of mitotic

activity and leukocyte infiltration. Luminal dilatation may per-sist into late estrus, although this is generally not the case.Some endometrial epithelial mitotic activity returns by the endof estrus.


FIGURE 1.—Diestrus. Black bars = 100 μm; red bars = 1000 μm.



Ovaries: Degenerate ovarian corpora lutea are often present atestrus, but others (the newly formed corpora lutea) are small,with cells showing a basophilic cytoplasm and occasionally acentral fluid-filled cavity retained from the follicular stage.Central fibrous tissue is generally not present.

Metestrus

Metestrus is illustrated in Figure 4.

Vagina: The start of metestrus is marked by the mid region ofthe vagina showing a complete detachment of the cornified


FIGURE 2.—Proestrus. Black bars = 100 μm; red bars = 1000 μm.



epithelium, generally with residual squames present in the lumen.As noted above, some cornified epithelium may persist, partic-ularly adjacent to the vaginal opening. There is a continued desqua-mation of epithelial cells throughout the stage, with a progressiveloss of the stratum granulosum and upper germinativum (stratum

spinosum). There is an accompanying variable leukocyte infil-tration. As noted under diestrus, the end of metestrus/start ofdiestrus is marked by the epithelium reaching its lowest level.The smear during this stage shows leukocytes, a few cornifiedcells, and basophilic cells.


FIGURE 3.—Estrus. Black bars = 100 μm; red bars = 1000 μm.



Uterus: During metestrus, the uterine endometrial epitheliumshows continued vacuolar degeneration, but also a marked returnof mitotic activity, so both are seen together. There is also avariable leukocyte infiltration

Ovaries: Ovarian corpora lutea may still contain a fluid-filledcentral cavity generally devoid of fibrous tissue, and the cyto-plasm of the new corpora lutea are somewhat less basophilicthan at estrus but smaller than that seen at diestrus.

THE CYCLE IN AGING RATS

During the midlife period in rats, the estrous cycle lengthincreases, and some histological inconsistency is occasionallyseen between the various components of the tract. Progesteroneplays an important role in influencing cycle length in rodents(Nequin et al. 1979), and progesterone and estrogen act syner-gistically (Morali and Beyer 1979). The estrogen/progesteroneratio is elevated for prolonged periods before ovulation in most


FIGURE 4.—Metestrus. Black bars = 100 μm; red bars = 1000 μm.



middle-aged rats, and this age-related deficit in progesteronehas a significant responsibility for this increase in cycle length(vom Saal et al. 1994). At any time from six to eighteenmonths, but seldom later, following lengthening of the cycle orinterspersed long cycles, a constant state of vaginal cornifica-tion or “persistent estrus” can be observed (vom Saal et al.1994). This may be followed by states in which cycles are tento fourteen days long, suggestive of a prolonged luteal phase,which is termed “repetitive pseudopregnancy” (Nelson andFelicio 1985). The final stage of reproductive senescence is“persistent anestrus” (vom Saal et al. 1994).

Persistent Estrus

Persistent estrus is illustrated in Figure 5.

Subsequent to the lengthening of the estrous cycle, mostaging females between six and eighteen months of age exhibitperiods of persistent estrus, with constant sexual receptivity(Felicio et al. 1984). This state results from estradiol levelsbeing maintained at a tonic level (Lu et al. 1979; Nelson et al.1981), reflecting the presence of numbers of ovarian follicularcysts (Gosden, Laing, Flurkey et al. 1983) and a tall columnarendometrial epithelium. The tonic estrogen secretion stimulates


FIGURE 5.—Persistent estrus and repetitive pseudopregnancy.



cornification of the vaginal epithelium, so this state is also referredto as persistent vaginal cornification (vom Saal et al. 1994). Lowlevels of progesterone during this period (Miller and Riegle 1980;Nelson et al. 1981) are considered responsible for the maintenanceof this state (vom Saal et al. 1994) and are associated with anabsence of corpora lutea (Gosden, Laing, Flurkey et al. 1983).

Repetitive Pseudopregnancy

Repetitive pseudopregnancy is illustrated in Figure 5.

In young adult female rats, whether sterile or fertile, for thefirst nine or ten days after mating, prolactin secreted by thepituitary maintains the corpora lutea. The placenta of a suc-cessfully mated female then takes over (Smith 1980; Smith et al.1975). If mating is unsuccessful, the corpora lutea degenerateby Day 12 or 13.

Therefore, the repetitive states of pseudopregnancy that canoccur following persistent estrus in aging rats are typically tento fourteen days in duration (vom Saal et al. 1994). This periodis characterized by maintenance of corpora lutea beyond thenormal two-day life span and is associated with high proges-terone levels (vom Saal et al. 1994). It is considered that thesechanges, following an elevated estradiol/progesterone ratioduring persistent estrus, result from reduced estradiol and ele-vated prolactin, often as a result of a pituitary tumor (Everett1980). The raised prolactin levels maintain persistence of thecorpora lutea. The vagina during this period shows a low stra-tum germinativum, but variable overlying mucification. Theuterus commonly exhibits a folded endometrial epithelium.

Persistent Anestrus

Figure 6 illustrates persistent anestrus.

Old rats eventually reach a final stage of reproductive senes-cence. This is termed persistent anestrus, or persistent diestrus.After the cessation of ovulation and entry into persistent estrus,there is still some maturation of follicles, followed by atresiawithout ovulation, culminating in complete follicular depletion(Gosden, Laing, Felicio at al. 1983; Gosden, Laing, Flurkey etal. 1983). Morphological features of the reproductive tract arecharacterized by a low, somewhat mucified vaginal epitheliumand a low inactive endometrial epithelium. The loss of folliclesresults in estradiol levels similar to those seen in ovariec-tomized rats (vom Saal et al. 1994). There are low progesteronelevels, reflecting the absence of corpora lutea (Lu et al. 1979),and high prolactin levels, generally from pituitary tumors,which have an inhibitory effect on LH (vom Saal et al. 1994).

REFERENCES

Advis, J. P., Andrews, W. W., and Ojeda, S. R. (1979). Changes in ovariansteroidal and prostaglandin E responsiveness to gonadotropins during theonset of puberty in the female rat. Endocrinology 104, 653–58.

Andrews, W. W., and Ojeda, S. R. (1981). A detailed analysis of the serum LHsecretory profiles of conscious free-moving female rats during the timeof puberty. Endocrinology 109, 2032–39.

Astwood, E. B. (1939). Changes in weight and water content of the uterus ofthe normal adult rat. Am J Physiol 126, 162–70

Blandau, R. J., Boling, J. L., and Young, W. C. (1941). The length of heat in thealbino rat as determined by copulatory response. Anat Rec 79, 453–63.

Everett, J. W. (1980). Reinstatement of estrous cycles in middle-aged sponta-neously persistent estrous rats: Importance of circulating prolactin and


FIGURE 6.—Persistent anestrus. Black bars = 100 μm; red bars = 1000 μm.



the resulting facilitative action of progesterone. Endocrinology 106,1691–96.

Felicio, L. S., Nelson, J. F., and Finch, C. E. (1984). Longitudinal studies ofestrous cyclicity in ageing C57BL/6J mice . II. Cessation of cyclicity andthe duration of persistent vaginal cornification. Biol Reprod 31, 446–53.

Freeman, M. E. (1994). The neuroendocrine control of the ovarian cycle of therat. In The physiology of Reproduction, 2nd ed. (E. Knobil, and J. D.Neill, eds.), Raven Press, New York.

Gosden, R. G., Laing, S. C., Felicio, L. S., Nelson, J. F., and Finch, C. E.(1983). Imminent oocyte exhaustion and reduced follicular recruitmentmark the transition to acyclicity in aging C57BL/6J mice. Biol Reprod28, 255–60.

Gosden, R. G., Laing, S. C., Flurkey, K., and Finch, C. E. (1983). Graafian fol-licle growth and replacement in anovulatory ovaries of ageing C57BL/6Jmice. J Reprod Fertil 69, 453–62.

Hartman, C. G. (1944). Some new observations on the vaginal smear of the rat.Yale J Biol Med 17, 99–112.

Heape, W. (1900). The “sexual season” of mammals and the relation of the“pro-oestrum” to menstruation. Q J Microsc Sci 44, 1–70.

Hubscher, C. H., Brooks, D. L. and Johnson, J. R. (2005). A quantitativemethod for assessing stages of the rat estrous cycle. Biotech Histochem80, 79–87.

Knobil, E., and Neil, J. D., eds. (1994). The Physiology of Reproduction, 2nded. Raven Press, New York.

Long, J. A., and Evans, H. M. (1922). The oestrous cycle in the rat and its asso-ciated phenomena. Mem Univ Calif 6, 1–148.

Lu, K. H., Hopper, B. R., Vargo, T. M., and Yen, S. S. (1979). Chronologicalchanges in sex steroid, gonadotropin, and prolactin secretion in ageingfemale rats displaying different reproductive states. Biol Reprod 21,193–203.

Maeda, K., Ohkura, S., and Tsukamura, H. (2000) Physiology of reproduction.In The Handbook of Experimental Animals—The Laboratory Rat. (G. J.Krinke, ed.), Academic Press, London.

Mandl, A. M. (1951). The phases of the estrous cycle in the adult white rat. JExp Biol 28, 576–84.

Parkes, A. S., ed. Marshall’s Physiology of Reproduction (1960). Longmans,Green, and Co., London, New York, Toronto.

Miller, A. E., and Riegle, G. D. (1980). Temporal chnges in serum progesteronein aging female rats. Endocrinology 106, 1579–83.

Morali, G., and Beyer, C. (1979). Neuroendocrine control of mammalianestrous behaviour, In: Endocrine control of sexual behaviour (C. Beyer,ed.), pp. 33–75, Raven Press, New York.

Nequin, L. G., Alvarez, J., and Schwartz, N. B. (1979). Measurement of serumsteroid and gonadotropin levels and uterine and ovarian variables through-out 4 day and 5 day estrous cycles in the rat. Biol Reprod 20, 659–70.

Nelson, J. F., and Felicio, L. S. (1985). Reproductive aging in the female: Anetiological perspective. Rev Biol Res Aging 2, 251–314.

Nelson, J. F., Felicio, L. S., Osterburg, H. H., and Finch, C. E. (1981). Alteredprofiles of estradiol and progesterone associated with prolonged estrouscycles and persistent vaginal cornification in aging C57bl/6J mice. BiolReprod 24, 784–94.

Ojeda, S. R., and Urbanski, H. F. (1994). Puberty in the rat. In The Physiologyof Reproduction, 2nd ed. (E. Knobil and J. D. Neill, eds.), pp. 363–409,Raven Press, New York.

Ojeda, S. R., Wheaton, J. E., Jameson, H. E., and McCann, S. M. (1976). Theonset of puberty in the female rat. I. Changes in plasma prolactin,gonadotropins, LHRH levels and hypothalamic LHRH content.Endocrinology 98, 630–38.

Smith, M. S. (1980). Role of prolactin in regulating gonadotropin secretion andgonad function in female rats. Fed Proc 39, 2571–76.

Smith, M. S., Freeman, M. E., and Neill, J. D. (1975). The control of proges-terone secretion during the estrous cycle and early pseudopregnancy inthe rat: Prolactin, gonadotropin and steroid levels associated with rescueof corpus luteum of pseudopregnancy. Endocrinology 96, 219–26.

Urbanski, H. F., and Ojeda, S. R. (1985). The juvenile-peripubertal transitionperiod in the female rat: Establishment of a diurnal pattern of pulsatileluteinizing hormone secretion. Endocrinology 117, 644–49.

vom Saal, F. S., Finch, C. E., and Nelson, J. F. (1994). Natural history and mech-anism of reproductive aging in humans, laboratory rodents, and otherselected vertebrates. In The Physiology of Reproduction, 2nd ed. (Knobil,E., and Neil, J. D., eds.), pp. 1213-1314, Raven Press, New York.

Yuan, Y., and Foley, G. L. (2002). Female reproductive system. In Handbook ofToxicologic Pathology, 2nd ed., Vol. 2. (W. M. Haschek, C. G. Rousseaux,and M. S. Wallig, eds.), pp. 847–94, Academic Press, London.




guide histologi vaginal smear

Documents