Maudsley Hospital, London, SE 5 occur in the blinded (Bissonnette, 1935, 1938)

17 Pages·2006·3.68 MB·English

102
J. Physiol. (I956) I3I, 102-II4
THE EFFECT OF PITUITARY STALK SECTION ON LIGHT-
INDUCED OESTRUS IN THE FERRET
BY B. T. DONOVAN* AND G. W. HARRIS
From the Department ofNeuroendocrinology, Institute ofPsychiatry,
Maudsley Hospital, London, S.E. 5
(Received 18 May 1955)
Exposure ofnormal anoestrous ferrets to extra (artificial) illumination during
winter induces an oestrous state (Bissonnette, 1932). This reaction does not
occur intheblinded (Bissonnette, 1935, 1938), orhypophysectomized (Hill &
Parkes, 1933) animal. From this and other data it is clear that the stimulus
oflight acts via the retina and optic nerve fibres to excite, in some way, the
secretion offollicle-stimulating hormone (FSH) from the anterior lobe of the
pituitary gland.
Several attempts have been made to determine the anatomical pathway
involved between the optic nerves and the pars distalis of the hypophysis
(Clark, McKeown & Zuckerman, 1939; Thomson, 1951, 1954). On general
grounds (Harris, 1948) it seems likely that the hypothalamus and pituitary
stalk would form part ofthis path. However, there are data contrary to this
view. Burger (1949) mentions that the sexual activation induced by artificial
long days was not lost in the stalk-sectioned ferrets studied by Bissonnette
(1938), but gives no information concerning post-mortem studies. Thomson &
Zuckerman (1953, 1954) have recently reported that complete interruption of
the pituitary stalk inthe ferret does not necessarily prevent the occurrence of
oestrus following extra illumination in winter.
Ifthe hypothalamus and pituitary stalk areimplicated inthelight-induced
oestrousresponse oftheferret, itisprobable thatitisthevascular component
ofthe stalk, viz. the hypophysialportalvessels, whichisresponsiblefortrans-
mitting the stimulus to anterior pituitary tissue (see Harris, 1948; Harris &
Jacobsohn, 1952). Since these vessels are known to regenerate in a large
proportion ofanimals after simplestalksection, and since validprocedures to
control anysuch regeneration do not seemto havebeenundertakeninstudies
* BeitMemorialResearch Fellow.
OESTRUS AND THE PITUITARY STALK 103
of the light response of the stalk-sectioned ferret, the problem was re-in-
vestigated. Thepresentresults do not confirmthose ofThomson & Zuckerman
(1953, 1954).
METHODS
Female albino ferrets, of280-780g body weight, were fed on a controlled diet ofbread, meat,
milk,yeast,cod-liveroilandasaltmixture.Threegroupsofanimalswerestudied: (a)fouranimals
inwhichthepituitarystalkwasexposedbutnotdamaged; (b) sixanimals in which thepituitary
stalkwasexposedandcut; and(c) fourteen animalsinwhichthepituitarystalkwasexposed,cut,
andarelativelylargewaxedpaperplateinsertedbetweenthepituitaryglandandhypothalamus.
After operation these animals were exposed to extra illumination during the winter months of
1953-4 and the development ofany oestrous response observed.
Operative technique. The pituitary stalk was exposed by a temporal route. Under anaesthesia
withpentobarbitone sodium (36 mg/kgbodywt., i.P.) theanimalwasimmobilizedonitsleftside
onananimalboard.Thetemporalismusclewasfullyexposedbyaverticalskinincisionmadefrom
the mid-line ofthe head superiorly to the posterior end ofthe zygomatic arch inferiorly (Text-
fig. Ia). AT-shapedincisioninthetemporalismuscleenabledittobesplit,andthetwohalvesto
be separated from the skull and retracted anteriorly and posteriorly (Text-fig. 1b). A large
areaofskullwasthenremoved (Text-fig. 1c), care being taken to leave the dura intact andto
controlanyoozingof blood with bone wax. The dura was next cut, so as to provide flaps to
cover the margins ofthe bone. The further stages in the operation were performed with the
aid ofahead lamp and with the ferret placed in an inverted position, thus utilizing theeffect
ofgravityinseparatingthebrainfromthebaseoftheskull. Afterslowretractionofthetemporal
lobe ofthe brain a wide view ofthe hypothalamo-hypophysial region was obtained, the main
featuresofwhichareshown in Text-fig. ld. Whenthehypothalamus, pituitary stalk andgland,
andsurroundingstructureswereallclearlyvisible,thestalkwastransectedwiththeaidofasmall
hookorknife. Inanimalsinwhichtheoperationofsimplestalksectionwasperformed,thecom-
pletenessoftheoperationwascontrolledbysweepingabluntprobewidelyacrosstheuppersurface
ofthe pituitary gland and observing the two separately movable ends ofthe stalk, by passing
cotton-wool pledgelets over the gland and removing them between the two internal carotid
arteries,orbyinsertingalargepaperplatebetweenthehypothalamusandpituitaryglandandthen
removingtheplate.Thewaxedpaperplatesusedinsomeoftheseanimals,andinothersinwhich
theplates wereleft permanently inposition, wereinserted first betweentheoptic nerves and
skullbase,inapositionanteriortotheinternalcarotidarteries.Theplateswerethengentlypushed
inacaudaldirection,betweenthearteries,sothatabouttwo-thirdsofthe platewas situatedin a
planeposteriortotheinternalcarotids.Thedimensionsoftheseplates(5x3mm)weresuchthat
theyalmostcompletelyspannedthedistance betweenthetwoarteriesandformedawidebarrier
between thehypothalamus and pituitary gland (seeP1. 1, figs. 1-4, andP1. 4, figs. 14, 16). The
operation was completed by replacing the dural flaps over the brain, suturing the temporalis
muscletocovertheexposedduraandbrainandclosingtheskinwoundwithinterruptedsutures.
Injections of40,000 units penicillin and 50mg streptomycin ('Seclomycin', Glaxo Laboratories
Ltd.) were given at operation and occasionally onthefollowing day. Recoverywasuneventful
and no disabilities were observed. At eventual autopsy the wound was found to be well and
cleanlyhealedinallcases. Someadhesionofthebraintooverlyingmuscleortotheskullfloorwas
occasionallyseen, aswasslight dilatation ofthelateralventricle ontheoperatedside.
Lighting conditions. The animals, in individual cages, were kept under general animal house
conditionsuntil26 October 1953 whentheywereallobservedto haveshrunkenvulvae. Atthis
timetheywereplacedundertheexperimentalconditionsofprolongedillumination. Eachanimal
wasplacedinashadow-lesscageonarack,andwasilluminatedsolelybya40WMazdaDaylight
TypeMCF/Ufluorescent lamp (Hart, 1951), placed at adistance of67cm from thefloorofthe
cage(P1. 2,fig. 5).Thelightingwasautomaticallyswitchedonandoffat8.00a.m. andmidnight
respectively.Thecagescontainedashallowlayerofsawdustorwoodshavingsandwerescreened
104 B. T. DONOVAN AND G. W. HARRIS
one from the other. The ferrets in the three different groups were distributed at random in the
racks. Eachanimalwasweighedandexamined,andtheconditionofitsvulvaandeyesnoted,at
weekly intervals. No case ofcataractwasobserved.
Autopsyandhistologicalprocedures.Thefirstanimalwaskilledon13January1954andthelast
on25February1954. Fourhoursbeforedeatheachanimalwasinjectedsubcutaneouslywith3,uc
of1311, andplacedinaglasstankinordertocollectanyexereta. Afterkillingwithchloroforma
sampleofbloodwasdrawnfromtheinferiorvenacavaintoasyringecontainingasmallquantity
ofsolidheparin, anyurineinthe bladderwas collected, andthe ovaries, reproductive tractand
adrenalglandsremoved. Thevascularsystemwasthenperfusedwithatleast200ml. ofdiluted
Mandarin Black indian ink (Winsor & Newton; 1 part indian ink to 3 parts distilled water),
injectedtowardtheheartthroughtheabdominalaorta. Afterremovalofthethyroidgland,the
thyroid, adrenals, ovaries and uterus were freed from extraneous tissue and weighed. These
tissues, except for one lobe of the thyroid gland which was weighed separately and used for
radioactive assay, were fixed in Zenker's solution, dehydrated, embedded in paraffin wax and
II
O.TX
c a
Text-fig.1.Illustratingthetemporalapproach.Thediagramsshow:(a)Themethodofimmobilizing
theheadandtheskinincision.Thenoseispassedthroughawireloopintheoperatingboard
and the jaws secured with tape. (b) The exposure and incision in the temporalis muscle.
S.,sagittalcrestoftheskull. (c) Theexposureofthecerebralhemisphere.(Theflapsofdura
materarenotshown.) S.,sagittalcrestoftheskull; M.,temporalismuscle. (d)Theviewof
thehypothalamo-hypophysial region following inversion ofthe ferret andretraction ofthe
temporallobe. H., hypothalamus; I.C., internal carotid artery; IW.B., mammillary body;
O.N., oculomotor nerve; O.T., optic tract; P.G., pituitary gland; P.S., pituitary stalk;
B.,retractor; T.C.,tentorium cerebelli.
OESTRUS AND THE PITUITARY STALK 105
sectioned at a thickness of10 . The sections were stained with haematoxylin andeosin. After
careful examination ofthe operation site and removal ofthe calvarium, the head was fixed in
10%formalin, decalcifiedinamixtureof40%formicacidand7%sodiumformate (Engelbreth-
Holm & Plum, 1951) and a block oftissue comprising the hypothalamic region and skull base
dehydrated,embeddedinlowviscositynitrocelluloseandseriallysectionedinthesagitalplaneat
a thickness of130-200u. Indian ink was not perfused through the vascular system oftwo an-
oestrous ferrets (56 and 59), but instead the heads were dissected and theposition oftheplates*
directly observed. The hypophyses were fixed in situ with Susa's solution, and serial sections
stained bythe Heidenhain-Azan technique.
Treatment ofradioactive samples. One lobe ofeach thyroid gland was dissolved in 2N-NaOH
andthevolumemadeupto3or6ml.withdistilledwater.Thebloodsampleswerecentrifugedand
the plasma separated. The urine samples were diluted to aknown volume with distilled water.
Theradioactivity ofthesolutions soobtainedwas measured, usingaLabgearscalerandanend-
windowcounter. Twoaliquotsofeachsolutionwerecountedandaftercorrectionforisotopedecay
thecountswereconvertedintomicrocuriesbymeansofacalibrationcurveobtainedfrommeasure-
ments ofthe activity ofthe originalsample ofradioactive iodine.
Measurement of 'pituitary volume'. The relative volumes ofthe pituitary glands, and ofthe
anteriorandposteriorlobes,weremeasuredinthedifferentanimalsbythe'paperweight'method.
Theimage ofthehypophysisineach section ofaseries wasprojected ontoasheetofpaperand
the outlines traced. The outlines ofeach series from apituitary gland were accommodated on a
singlesheetofpaper,andeachserieswastracedthreetimesusingthreeseparatesheets. Bycutting
outthedemarcatedareasofpaper,andweighingthem,figureswereobtainedrelatedtothevolume
ofpituitarytissue,andofthetwolobes,inindividualglands. Theweightsforeachgland, andthe
average valuesfor the three groups ofanimals, are givenin Table 1.
Other observations. The water consumption of some ferrets was measured daily for a 16-day
period, with the intention ofdetermining whether the stalk-sectioned animals developed a state
ofdiabetes insipidus. Toseewhether ovulation wouldfollow copulation in thestalk-cutanimals
attempts were made to mate five ofthe oestrous females. Since males were not exposed to the
increasedillumination onlysexually quiescent maleswereavailable forthispurpose, andcopula-
tion was never observed to follow the sporadic sexual activity. Pairs were occasionally left
together overnight.
RESULTS
Development ofoestrus
Twelve ferrets out of the twenty-four developed an oestrous state, as judged
by vulval swelling, within 2-7 weeks from the start of exposure to increased
illumination. The other twelve animals remained anoestrous till killed after
14-18 weeks of exposure. As is shown in Text-fig. 2, all four blank-operated
ferrets, five ofthe six simple stalk-sectioned animals and three ofthe fourteen
ferrets with paper plates inserted became oestrous.
Hypothalamo-hypophysial region
Blank-operated animals. The four animals in which the pituitary stalk was
exposed but not damaged provided a basis of comparison for the stalk-cut
animals. Thetypicalappearance ofthe region maybe seen in P1. 3, fig. 6. Itis
evident that the ferret possesses a poorly developed primary plexus, and the
capillary loops passing up into the median eminence are few. However, well-
marked portal vessels run caudally along the stalk to enter the anterior pole
106 B. T. DONOVAN AND G. W. HARRIS
of the gland. The normal infundibular process (posterior lobe) is large anda
prominent cleft separates this structure from the pars distalis.
Simple stalk-sectioned animals. Five of this of six ferrets became
group
oestrousandinallfivesubstantialvesselswereobservedtoconnectthemedian
eminence ofthe-tuber cinereum and the distalis of the pituitary (P1. 3,
pars
fig. 8). Theneurallobeofthepituitaryintheseanimals markedlyatrophic,
was
Weeks under
'long day' 2 3 4 5 6 7 8 9 10 11 12 13 14
conditions
28
34
. 38
C 39
V40
-~46
51
V)
52
55
56
59
26 _ _ _ _
(4)
0~
Text-fig. 2. Illustrating theonsetofoestrusinthethreegroups ofanimals.
and found to be vascularized only in the central region. The pituitary stalk
oftenappeared swollen at the site ofstalk section, possibly dueto scarforma-
tion though special neuroglial stains were not used to investigate this point.
The pars distalis was in all cases seen to be well supplied with blood vessels
which formed a dense plexus throughout this part ofthe gland.
Sectionsthroughthehypothalamicregionofoneanimalinthis group (ferret
26)whichfailedtobecomeoestrousrevealedveryfew,ifany,vesselsconnecting
OESTRUS AND THE PITUITARY STALK 107
the well-marked vascular fields of the median eminence and the anterior
pituitary gland (P1. 3, fig. 10).
Stalk-sectioned animals with plates. Three of the fourteen animals in which
the pituitary stalk had been cut and a paper plate inserted between the stalk
endsbecameoestrous. Ineachofthesethreeanimalsmicroscopic examination
'showed that the plate had been misplaced (in two being too far forward
(P1. 4, fig. 12) and inone beingplacedobliquely) andthatvessels werepresent
connectingthemedianeminenceandtheparsdistalis aroundtheborderofthe
plate.
Eleven animals in this group remained anoestrous for the duration of the
experiment. In two of these ferrets the position of the paper plate was in-
vestigated by dissection only, and in both cases the plate was found to be
situatedsquarelyoverthepituitaryglandandsealedtotheskullfloorbyfibrous
tissue (P1. I,figs. 1, 3). Intheremainingnineanimalsmicroscopicexamination
showedtheplatesalsowellsituated. Ineachcaseitintervened overawidearea
between the hypothalamus and pituitary gland, forming an effective barrier
to regeneration ofthe hypophysial portal vessels (P1. 4, figs. 14, 16). The only
possible vascular connexions between the median eminence and the anterior
pituitary gland consisted ofa few fine capillaries runningin the fibrous tissue
surrounding the plate (as shown in P1. 4, fig. 14).
In all animals in this group injected with indian ink andexamined micro-
scopically, the median eminence contained a prominent primary plexus, the
anterior lobe was well vascularized and the posterior lobe was atrophic and
vascularized only in its central zone.
Volume ofthe hypophysis
The results of the measurements of the relative hypophysial volumes are
giveninTable 1. Thesedatashowthata markedatrophyoftheposteriorlobe,
and a slight reduction in size ofthe anterior lobe, follows stalk section in the
ferret. It may be noted that the volume ofanterior pituitary tissue present
in the animals that remained anoestrous appears to be greater than in the
animals thatdevelopedoestrus. Thisisprobably withoutsignificance sincethe
volume of posterior lobe tissue in the two groups is in a similar proportion.
Ovaries
Theovariesoftheoestrousand anoestrousferretsshowedmarkeddifferences
both in weight (Table 2) and in macroscopic and microscopic appearance. In
the oestrous group the ovaries showed large follicles. One of the oestrous
females (ferret 21), placed with a male, was found to have ovulated. None of
the ovaries from the other animals showed luteinization. The ovaries of the
anoestrousferrets were small, atrophic structures which containedprimordial,
108 B. T. DONOVAN AND G. W. HARRIS
but no antral, follicles. The ovaries ofa normal anoestrous ferret, killed with
the operated anoestrous group, were large (weighing 146 mg) in comparison
with the stalk-sectioned anoestrous females and contained numerous small
follicles with antra.
TABLE 1. Therelativevolumes, asdetermined bythe'paperweight' method, ofthe
pituitaryglands ofthe controlandexperimental animals
Thefiguresin brackets expressthe volume of theglands and the lobes as a percentage of
those oftheblank-operated controls
No. of Pituitary Anterior Posterior
animal gland lobe lobe
Blank-operated 44 87 53 34
controls 48 66 32 34
54 71 39 32
58 61 37 24
Average 71 (100%) 40 (100%) 31 (100%)
Experimental 14 33 24 9
oestrous animals 21 46 34 12
22 55 40 15
31 40 30 10
32 39 28 11
35 33 25 8
41 47 36 11
43 54 36 18
Average 43 (61%) 32 (80%) 12 (39%)
Experimental 26 56 46 10
anoestrous 28 51 38 13
animals 34 52 42 10
38 43 32 11
39 55 36 19
40 55 38 17
46 44 32 12
51 41 27 14
52 50 34 16
55 76 55 21
Average 52 (73%) 38 (95%) 14 (45%)
Adrenals
As may be seen in Table 2 only a slight reduction in weight ofthe adrenal
glandsoccurredintheanoestrousgroup. Calculation, usingthe't'test,showed
thedifference inabsoluteadrenalweightsoftheoestrousandanoestrousstalk-
sectioned groupsto be significant (P=001). The differenceinadrenalweights
calculated on a body-weight basis is only probably significant (P=0.05).
Histological differences were not observed in the haematoxylin and eosin
stained sections ofthe adrenal glands ofthe two groups.
Thyroidglands
The difference inthemeanweightsofthethyroidglandsoftheoestrousand
anoestrous groups wasnotstatisticallysignificant. Microscopicallytheglands
of the anoestrous animals appeared to have flatter epithelium and more
colloid than those in the oestrous group, but this observation is rendered
OESTRUS AND THE PITUITARY STALK 109
doubtful by the great variation in the microscopic picture found in different
areasofanyonegland. Thefiguresobtainedforthe4 hruptakeofradio-iodine,
however, show a clear difference betweenthe oestrous andanoestrousanimals
(Table 2), and indicate a decreased activity of the thyroid gland in the
anoestrous group. A study ofthe plasma concentration of1311 at the time of
TABLE 2. Organ weights andthyroidal131Iuptake ofcontrol and
experimental animals
Thyroid
131Iuptake
%inj.
Body Ovaries Uterus Adrenals dose/
Ferret wt. (mg/kg (mg/kg (mg/kg (mg/kg 100mg
no. (g) bodywt.) bodywt.) body wt.) bodywt.) gland
Blank-operated 44 1330 126 652 115 44 19
animals 48 660 171 1318 144 88 24
54 600 142 1415 226 96 19
58 880 122 1370 127 60 14
Average 867 140 1189 153 72 19
Experimental 14 750 141 1264 121 61 22
oestrous animals 21* 760 189 2426 147 106 21
22 860 88 737 152 73 10
31 610 184 1413 187 102 14
32 780 104 1332 146 92 6
35 840 145 1017 125 64 21
41 710 118 1251 155 73 9
43 1000 130 931 155 50 24
Average 789 137 1296 148 78 16
Experimental 26 580 36 426 155 109 7
anoestrous 28 680 34 390 129 119 6
animals 34 1160 15 90 116 84 5
38 610 31 131 93 103 6
39 760 41 125 135 55 7
40 840 30 127 98 102 4
46 840 26 108 86 80 8
51 890 31 111 99 75 4
52 800 26 134 111 75 6
55 680 40 132 143 57 9
56 660 61 136 145 91 9
59 640 58 127 177 83 9
Average 762 36 170 124 86 7
* Thisanimalwaspseudopregnant.
death, andoftherenal excretion of1311inthe4 hrpriorto death, showedthat
changes inthe fluid space ofthe bodyorin kidneyfunction could not explain
thedifferencesinthyroidal-uptakeof131Jobservedinthetwogroupsofanimals.
Body weight
Little difference was observed in the average body weights of the oestrous
and anoestrous ferrets. It was noticed, however, that the weights of the
oestrous ferrets tended to fallwhen they came into heat. For example, ferret
43weighed1120 gatthetimeartificialilluminationstarted, 1460g6weekslater
110 B. T. DONOVAN AND G. W. HARRIS
whenitfirst showed signs ofoestrus, and only 1000 g whenit was killed after
8 weeks in heat. The anoestrous ferrets showed either a relatively constant
weight or a gradual increase in weight.
Water consumption
A pronounced polydipsia was not observed to follow stalk section in the
ferret. When measured over a period of 16 days the maximum daily con-
sumptionofwaterwasseeninferret22whoseaveragedailyintakewas 136ml.,
whilst two control animals drank an average of 50 and 56 ml. water daily.
DISCUSSION
In drawing conclusions regarding the function of the adenohypophysis from
the results ofexperiments involving pituitary stalk section attention must be
paid to the possibility that signs of anterior pituitary deficiency are due to
ischaemic necrosis of the gland. As pointed out by Greep & Barrnett (1951),
section ofthe pituitary stalk might cut offsufficient ofthe total blood supply
to the gland to result in atrophy ofanterior lobe tissue and a state of 'hypo-
physectomy'. In the present study it was found that 80-95% ofthe normal
volume ofanterior tissue was present in the stalk-sectioned animals, and that
the ferrets in which a lack ofpituitary response to light was observed had, if
anything,largerglandsthanthe stalk-sectionedanimals thatbecameoestrous.
Itwouldseem clearthattheloss ofthelight-induced oestrous response cannot
be attributed to an inadequacy in the total blood supply with consequent
atrophy ofthe gland.
The loss of the oestrous response following pituitary stalk section may be
mosteasilyexplainedasduetoseverance ofareflexpathinvolvedinpituitary
stimulation. The two components ofthe pituitary stalk that might form part
of such a path are the hypothalamo-hypophysial nerve tract and the hypo-
physial portal vessels. The evidence argues strongly against the possibility
that the nerve tract is involved since very few, if any, nerve fibres pass
to the pars distalis of the pituitary and, furthermore, this tract does not
regenerate if damaged so that uniform results would be expected to follow
stalksection. Ontheotherhand, thereisstrongevidencethatthehypophysial
portal vessels passing from the median eminence ofthe tuber cinereum to the
pars distalis of the gland are intimately concerned with maintaining and
regulating anterior pituitary activity (Harris & Jacobsohn, 1952). During
the present studyit was observed that regeneration ofthe hypophysial portal
vessels may follow section ofthe pituitary stalk in the ferret, thus confirming
similar observations made on the mouse (de Groot, 1952), rat (Harris, 1950),
ferret (Thomson & Zuckerman, 1953), rabbit (Jacobsohn, 1954) and monkey
(Harris & Johnson, 1950). Itwas also observed thatthe occurrence ofalight-
inducedoestrousresponseafteroperationcould becorrelatedwiththepresence
OESTRUS AND THE PITUITARY STALK ill
ofsuch regeneration. No animal in which regeneration ofthe vessels was pre-
vented came into heat, but vascular connexions across the site ofsectionwere
clearly observed in all eight stalk-cut animals that became oestrous. The con-
clusion is drawn that the hypothalamus and the hypophysial portal vessels
form part ofa neurovascular path transmitting the reflex stimulus, initiated
by light, to the adenohypophysis.
In analysing reports dealing with the effects of pituitary stalk section on
anterior pituitary function attention should be paid to the following technical
points:
(a) Operative route used to section the stalk. Two surgical approaches have
been mainly utilized. The buccal or parapharyngeal approach is easy to per-
form, it allows a clear view of the stalk but not of the general region, and
offerslittlescopeforplacingplates betweenthecutendsofthestalktoprevent
regeneration. Further disadvantages of the method are that any plates
manipulatedbetweenthehypothalamusandhypophysis tendto be eliminated
from the wound, and that the anterior pole of the pituitary gland tends to
prolapse into the trephine hole in the base ofthe skull (see photomicrographs
of Barrnett & Greep, 1951; Thomson & Zuckerman, 1954) so that the inter-
pretation ofhistological sections through the region is rendered difficult. The
alternative temporal approach may be, in certain forms, a more difficult
procedure, but it gives a clear view ofthe whole region and allows easyaccess
for placinglarge plates between the ends ofthe stalk. With this latter method
the field ofthe pituitary stalk is left clean and surrounded byintact meninges
and bone: plates inserted do not tend to become misplaced: and the pituitary
gland remains in its normal position. For these reasons the final anatomical
study of the region is facilitated.
(b) Methods employed to investigate the vascular archttecture of the region
following stalk section. Since the hypophysial portal vessels may regenerate
with speed and facility around, or through the interstices of, foreign bodies
(plates, pledgelets of wool) placed in their path, special attention should be
paid to the possibility of such regeneration. To ascertain the presence or
absenceofregeneratedvesselsitisnecessarytoperfusethearterialsystemwith
a sufficient quantity ofindian ink immediately after death, and to cut serial
sections at a thickness of 100-200 through a decalcified block oftissue con-
u
taining the base of the skull, pituitary gland and hypothalamus. Only with
the aid ofsuch thick sections may the vascular pattern be clearlyinterpreted.
In the reports of Thomson & Zuckerman (1953, 1954) it was claimed that
two ferrets in which hypophysial portal vessels had been completely inter-
rupted became oestrous when exposed to additional illumination during
winter. It is believed that the discrepancy between these results and those
observed in the present study may be adequately explained by the fact that
Thomson & Zuckerman used the parapharyngeal approach to the pituitary