Stimulation of the thyroid gland of a teleost fish, Gillichthys mirabilis, by tetrapod pituitary glycoprotein hormones

Coup. Bioc/a~. P/I.wo/. Vol. 72A. No. 3. PP. 477 to 482. 1982 Printed in Great Britain.

0300-9629/82/030477-06803.00/O 0 1982 Pergamon Press Ltd

STIMULATION OF THE THYROID GLAND OF A TELEOST FISH, GZLLICHTHYS MIRABILZS, BY TETRAPOD PITUITARY GLYCOPROTEIN HORMONES DUNCAN S. MACKENZIE* Department

of Zoology,

University (Received

of California. 13 Nooember

Berkeley,

CA 94720, U.S.A.

1981)

Abstract-l. Thyrotropins and gonadotropins purified from representatives of each tetrapod class and subunits of the mammalian hormones were tested for their ability to stimulate acute in t’iuo thyroxine release in a teleost fish. 2. All intact hormones showed significant, intrinsic thyrotropic activity in this fish; subunits were inactive. 3. Gonadotropins and thyrotropins capable of stimulating the teleost thyroid are therefore found in all tetrapod classes. 4. Independent evolution of teleost fish has apparently led to an unusual situation in which the thyroid receptors for glycoprotein hormones are unable to distinguish between these structurally similar molecules from heterologous species.

INTRODUCTION

When purified pituitary hormones first became available, Fontaine (1969) demonstrated that the teleost thyroid gland could be stimulated not only by mammalian and teleost thyrotropins. but by mammalian gonadotropins as well. In contrast, the mammalian thyroid showed strict specificity for mammalian thyrotropins. It has since been shown by other workers (Grau & Stetson, 1977, 1979; Mime & Leatherland, 1980) that there are many mammalian hormones which stimulate the teleost thyroid, including pituitary and placental gonadotropins and growth hormone. Fontaine called hormones such as these which only stimulate the thyroids of heterologous species “heterothyrotropins” and suggested that the common ability of the glycoprotein hormones to stimulate the teleost thyroid was due to their structural similarity, having evolved from a common, ancestral glycoprotein molecule (Fontaine, 1969; Fontaine & Burzawa-Gerard, 1977). On the basis of results with crude pituitary extracts, Fontaine (1969) also predicted that heterothyrotropins active in fish existed in other vertebrate classes. Until now, however, only hormones purified from the pituitaries of mammals and fish have been tested in teleosts; there is no information on the specificity of the teleost thyroid response to pituitary glycoprotein hormones from other classes. Chemical and biological evidence now exists indicating that these thyrotropins (TSH) and gonadotropins (follicle-stimulating hormone, FSH, and luteinizing hormone, LH) from other classes are also homologous to mammalian thyrotropins and gonadotropins (Licht er u/., 1977; Fontaine & Burzawa-Gerard, 1977). For this * Present address: Alberta. Edmonton, c .“.P.

72 3A

c

Department of Zoology, University Alberta T6G 2E9 Canada.

of 477

reason, one might expect that the receptor on the teleost thyroid. which interacts with teleost thyrotropins. mammalian thyrotropins and mammalian gonadotropins because of the structural similarity of these three molecules, would interact with the structurally similar pituitary glycoprotein hormones from other tetrapod species as well. The goal of the present study was to re-examine the phenomenon of teleost thyroid specificity in more detail. It is now possible to test various vertebrate thyrotropins and gonadotropins in a teleost system because these hormones are available in purified form from representatives of all tetrapod classes (Licht et al., 1977; MacKenzie, 1981; MacKenzie et al., 1978, 1981). In addition, it has been shown that thyroxine release may be a sensitive thyrotropin bioassay that can easily be performed in teleosts (Chan & Eales, 1976). In the present study, such a bioassay technique was used as a tool to study the hormonal specificity of thyroid response of another teleost. the goby Gillichthys mirabilis, to a diverse group of vertebrate pituitary glycoprotein hormones. This study provides an opportunity to confirm that a teleost fish responds to heterothyrotropins from all tetrapod classes. The response to these hormones will provide further evidence for their structural and functional similarity. MATERIALS AND METHODS Mature Gillichrh~s mirabilis which had been collected from San Francisco Bay at all times of the year were obtained from a commercial supplier and kept in artificial 100% sea water on a 12L:12D photoperiod for 46 weeks at 15°C. Animals were not fed during this time in an attempt to reduce intragroup variability (Ortman & Billig. 1966; Fontaine, 1969; Chan & Eales. 1976). Because preliminary experiments indicated that responses to bovine TSH were more variable in females than males. attempts were made to select males based on external appearance

lower J‘LU. r’cs~lt. XC OO”,,

(dongalcd

Cl~lng~ltcJ rlllUl p
of the animals used in thla ~tudq \b\cit’ males; wcipht, ranged from I I to 32f (L ~~16.61. Animal!, ,h

a

here removed

from the IS room and acclimated to room (X&26’ C) and natural photoperiod for 2 days prior to injection. Acclimated fish were randomly diwded into groups of six and each group was kept m aerated 100°10 sea water In a covcrcd plavtlc cage for the duration of the cxpcriment. Animals rccclvcd two hormone Injcctlons given IS hl apart. Hormones were lnjcctcd intraperitoneall] ~n a columc of 100 hiI with a 30 eauye needle rnscrted through the tall musculature to prevent leakage. At a prescribed time (see Results section) after the second InjectIon. blood was collected from the caudal \asculature Into heparimred hematocrit tubes. Plasma was removed after centrifugation and kept frozen for radioImmunoassay (RIA). RIA of thyroxine (T,) In unextracted plasma was performed b> the method of Chopra (1978) as prekiously described (MacKenzie er ul., 1978). and results of the T, RIA were analyzed on a DEC-20 computer wth a parallel-line RIA program. Analysis of variance and Duncan‘s multiple range test or Mann-Whitney II test for significance were also performed on the DEC-20 using the SPSS program package. Hormone potencies with 95”,, confidence limits (95”” cl.) were calculated using a computer program for parallel-line bioassays based on Finney ( 1964). Hormones were available from representatives of all tetrapod classes. All bioassays included at least 2 doses of NIH bovine TSH-B4 (2.12 l1.S.P. I! mg. supplied by the National Institutes of Health). A highly purified bovine TSH (approximately 5 10 U.mg) and its ii subunit were supplied by Dr J. Pierce. Ovine FSH (G4-21 IB). owne LH (G3-256DA). bovine LH (G3-235DAj and subunits of the ovine gonadotropins were prepared by Dr Harold Papkoff. as were all non-mammalian hormones. All three mammalian gonadotropins had IOU thyrotropic activity in a mammalian TSH-specific bioassay (all were less than 0.05 times the NIH TSH in the turtle radioIodIne uptake bloassay of MacKewe er u(.. 1978. 1981). Sea turtle TSH (TCX8B) and LH (TC75BRB) are described In MgcKenzie ef al. (1981); sea turtle LH is less than 6”,, as active as sea turtle TSH m a turtle TSH bioassay. Ostrich TSH (SCSBRC). LH lSC6BRB). and FSH (SC2OB) are described in MacKenzie (IY81): the ostrich gonadotropms are less than 3”,, as active as ostrich TSH in an avian TSH hioastemperature

Ld.._._

Fig. I. Time course

of bovine

_l_i___

2

4

TIME

AFTER

6

K&:41 I ‘I4

Inittal

experiments

the time the

course

T,-rclcasc

bovine

response

SECOND

undcrtakcn I-csponse of

lo detcrmtnc characteristics

Gi//ic~l~r/r~~.\to

the

of NIH

TSH

standard. Fqure 1 shows the response to doses of this bo\inc TSH and saline m In groups of animals ktllcd 4. X and IS hr after

4 diffcront May the

second

injection.

doses of bovine 4 hr:

this

.Thc

TSH

tish died in these groups. fish

in

two

clearly two

groups

distinct

to

the

to the fact that

3 and

the total

4.

The

doses

lower

(P

< 0.05)

same

doses

at 4 hr:

tno

doses still

than

show

T,

the levels le\cls

in

remain

number

waiving

the

levels in fish

;mimals

significnntlq receiving

the

recci\lng

the

at maximum

at X hr.

none

w~up c cantly

receiving

the highest

dose. hacl Tj

different

from

111saline-injcctcd

found

a similar

between

experiment. there

those

rcsponsc

2 and

X hr.

It was desirable

were distinct

to the increasing experiments sampling

~lth

were blood

controls. doses W;I~

rhc purposes

of thih

to choose ;I time at which bctv+een the rcsponscs

for this reason

conducted 6 7 hr

F3>

of the

lcvcls slgnili-

to increasing

diffcl-cnces doses and

the cxccption

For

(of arc

IX hr.

In August.

of the groups.

at

several

responses

bq X hr. Ichcn the ammals

TSH

different

indistinguishable

rcducmg

to

lower

hi&r

response

1~~ cwntially

may he due 111part

.d’ter

by klllrng the

all further

animals

second

and

hormone

injection.

Rqmw To

to r,~irrnr17ctlic11~/w~nor~c~.\ confirm

that

hetcrothyrotropic

~__L___L. ..I - -1 IO

12

INJECTION

14

mammalian in

LSollne

8

wcrc

and dose

16

gonadotropins

C;i//ic/~r/~~.\. both

ovine

1 18

(HOURS)

of T, release in G~llichthv.\ In response to mjectlon of saline or four Incr-caalng thyrotropin (NIH TSH-B4) standard. Values are means i_
doae\

uerc and

Gillichtl~~~s :hyroid

Dose

Fig. 2. In luteinizing

stimulation

(,ug)

rice

T, release response of Gillichrhys to bovine thyrotropin standard (NIH bTSH). ovine hormone (oLH). and ovine follicle-stimulating hormone (oFSH). Values are means f stan-

dard errors.

greater relative response to the NIH bovine TSII standard, whereas the heterothyrotropic response to both ovine gonadotropins (each tested at one dose) remained approximately the same. Bovine LH also demonstrated significant heterothyrotropic activity at the one dose tested. The highly purified bovine TSH was approximately twice as potent as the NIH TSH. In contrast to the intact hormones, subunits showed negligible thyrotropic activity. The potency of the bovine TSH /I subunit was less than lo”;, that of the highly purified bovine TSH (potency vs NIH TSH standard = 0.15. 957” c.1. = 0.04-0.52). This value is similar to the potency of this bovine TSH in a bioas-

bovine hormones were tested for their ability to stimulate T, release in this fish. In addition, to characterize further the nature of the specificity of the thyroid response, subunits of both ovine gonadotropins and bovine TSH were tested for activity. Figure 2 shows that in an assay in late June, the response to both ovine gonadotropins at two doses is essentially indistinguishable from the response to the NIH bovine TSH standard, although accurate potency estimates cannot be calculated because the responses to the lower doses of the ovine gonadotropins are not different from those in saline-injected controls. Figure 3 shows that in an assay in mid-August, there was a

701

I 601

/ / /hbTSHP IO

a c

/

WIIX

/ -Z-Z-;-_:--:<-

oFSHP s#oFSHa &,LHa \oLHP

01

05

10

Dose

50

100

(,ug1

Fig. 3. In rice T, release response of Gillichf/l~,s to bovine thyrotropin standard (NIH bTSH). highly purified bovine thyrotropin (bTSH), and its b subunit (bTSH/Q bovine luteinizing hormone (bLH). and ovine gonadotropins and their x and p subunits (oLH. oLHa. oLHfi. oFSH, oFSHr. oFSHP). Values are means f s!andard errors.

5 IO y Nltl

Hormone

TSH-B-l

I i v NIH TSWB4

Saline NIH bovine TSH

Sea turtle LH

i 75”,, NIH TSH-B4

TSH

c S”,, NIH TSH-B4

SW turtle TSf-l. LH O\II-ICII FSH f3v\Inc TSfj. 1.H O\lnc FSf1. LH O\t~-tch LH. TSH Bullfrog TSH Bullfrog l-t-1 Hov~neTSff /i Ovtnc FSH and LH subunits

Bullfrog LH TSH. LH

say specific for mammalian thyrotropin (potency m the turtle radioiodine uptake assay was approximately 0.10 times the NIH TSH). The subunits of the gonadotropins had no thyrotropic activity whatsoever at the doses tested. Response to non-mummalicm horrnonc~~~ Difficulties were encountered with the various nonmammalian preparations because of fluctuations in the response to these hormones and the NIH TSH standard. All hormones were tested in at least two assays. and attempts were made to choose doses which would give a linear doseeresponsc parallel to that of the standard. Unfortunately. because of variability in the assay, this was not always possible; the sensitivity of Gi/lichth~s to the various hormone preparations changed so dramatically over the course of the four months in which these assays were performed (September-December) that it was difficult to predict with accuracy the range of doses giving a linear response. For this reason. it was impossible to determine with precision the exact potencies of many of the various non-mammalian hormone preparations, and data were considered primarily on qualitative grounds to establish which fractions wcrc the most potent. The data in Tables 1 and 2 were chosen to illustrate the differences in potencies of the non-mammalian hormones. These data reflect the general

Table

2. Thyroxine

Hormone Saline NIH bovine TSH

Ostrich TSH/LH LH FSH

rclcasc ostrich

111 GI//~c,/I~/II.\ in response hormones

Dose (pg fish)

Y plasma T, (ng ml * S.E.M.)

0.5 1.5 4.5

9.X 31.1 65.4 74.2

i F k f

2.8 6.4 8.7 21.5

0.17 0.5 I .o 3.0 0.1 0.3

35.6 * 13.3 76.6 * 10.4 TX.4& 15.7 Xl.‘) * 16.7 57 5 + 17.5 75.5 + 17.1

to

trends in potencies seen over three assays and summarized in Table 3. Bullfrog hormones gave the most consistent response of any of the non-mammalian hormones (Table 1). Both bullfrog LH and TSH possessed significant activity in the two assays in which they were tested. The LH-contaminated TSH (C195B) and the NIH bovine TSH standard were equipotent in both assays (potency of C195B = 1.1 x NIH TSH-B4, 95”,, cl. = 0.48-2.69). whereas the bullfrog LH was 25”,,, as active (potency of Cl53C = 0.25 x NIH TSH-B4. 95”,, cl. = 0.14-0.46). The sea turtle was the only non-mammalian species from which pituitary hormones free of significant cross-contamination were available (MacKenzie et rrl.. 1981). Both sea turtle TSH and sea turtle LH were tested in three assays, and in all three both hormones were highly potent. In each case, however, these preparations either gave maximal stimulation. or were not tested at doses which gave a parallel response to the standard. Table 1 shows a typical response to these two preparations. There was no significant difference between the activity of the two in any assay. and they were usually 5-10 times more potent than the NIH bovine TSH standard. An example of typical results with the ostrich hormones is shown in Table 2. Of the three hormones tested, ostrich FSH was usually the most potent. although it was once again difficult to calculate exact potencies due to non-parallelism with standards. In the two assays in which it was tested, ostrich FSH was approximately 10 times more potent than the NIH bovine TSH standard. Ostrich TSH was approximately 50”,, as potent as the ostrich FSH in the same two assays. but response to the TSH was parallel to the response to the NIH bovine TSH standard. with a potency of 3.7 (95”,, cl. = 2.1-6.5). Ostrich LH was roughly equipotent to the NIH bovine TSH in both assays in which it was tested (potency = 0.81 x NIH TSH-BA 95”” cl. = 0.38-1.8). A summary of the approximate potencies of all these preparations appears in Table 3. DISCUSSION

It has been proposed that in tits release of thyroxine in teleosts can be used as an accurate bioassay of exogenously-administered thyrotropin (Chan & Eales. 1976). Ip the present study, this technique has been adapted to an estuarine goby, Gillichth~s miruhi/is, for the purpose of studying the response of this

Gillichth~s

thyroid

fish to a variety of tetrapod pituitary glycoprotein hormones. The sensitivity of the present assay (approximately I mIU/fish) is considerably better than that of other fish thyrotropin bioassays (cf. Table 3 in Chan & Eales, 1976), and the time course of response differs from that found in trout by Chan & Eales. The prolonged elevation of plasma T, (l&SO hr) seen in trout after 1 injection of thyrotropin is not seen in Gillichthys. The lack of a prolonged elevation of T, in Gillichth~s may be the result of intraperitoneal injection (versus intramuscular injection in trout) or the temperature at which they were tested. No attempts were made in the present study to determine the effects of temperature on the thyroid response, and fluctuations in room temperature (20-26’) may have contributed to the variability of the assay. Interassay variability may also be due to a seasonal change in thyroid sensitivity and magnitude of response; such a seasonal change has been reported for other teleosts (Grau & Stetson, 1978). Assay variability meant that the heterothyrotropic activity of the various tetrapod hormone preparations frequently could not be accurately determined in a quantitative manner. However, the qualitative relationships of these preparations to one another and to the NIH bovine TSH standard were reasonably consistent from one assay to the next, and were adequate to establish whether the various gonadotropins possessed heterothyrotropic activity. In general, if a particular gonadotropin demonstrated greater thyrotropic activity than could be accounted for by its thyrotropin contamination (as determined during hormone purification), the gonadotropin could be considered to possess heterothyrotropic activity in the fish. Using this criterion, Table 3 demonstrates that all gonadotropins tested possess heterothyrotropic activity in Gillichthys. All three intact mammalian gonadotropins are far more potent in the fish (l-4 U TSH activity/mg) than they are in bioassays specific for mammalian thyrotropin (less than 0.1 U TSH activity/mg). Gillichth.~ therefore resembles other teleosts in showing a heterothyrotropic response to intact mammalian gonadotropins. The results with the subunits of the mammalian hormones indicate, however, that the thyroid response in this teleost does possess a degree of specificity. Although these subunits, which can be reassociated to give full gonadotropic activity, must closely resemble the intact gonadotropins in many respects, they are inactive at the doses tested, with the exception of the bovine TSH fi. Because the activity of this particular subunit is similar in a thyrotropin-specific bioassay, however, its activity here is probably due to contamination with intact thyrotropin. Fontaine-Bertrand et al. (1981) have shown that subunits of bovine gonadotropins can be more potent than intact bovine gonadotropins in stimulating cyclic AMP accumulation in the teleost gonad, but a similar situation does not exist with the teleost thyroid, as intact molecules are required for full heterothyrotropic activity. Heterothyrotropins also exist in all other tetrapod classes. All three ostrich hormones possess significant activity in Gillichtkys and both ostrich gonadotropins, which were less than 3”; as active as the ostrich thyrotropin in an avian bioassay, are now as potent as ostrich thyrotropin in stimulating the fish thyroid.

stimulation

481

Ostrich FSH is, in fact, among the most potent of the hormones tested in this fish. In addition, both the sea turtle thyrotropin and LH are extremely potent, joining ostrich FSH as the most potent hormones tested in Gillichthys. Sea turtle LH is ten times more potent in relation to sea turtle thyrotropin in this fish than it is in a turtle thyrotropin bioassay (MacKenzie er al., 1981). Finally, although the bullfrog hormones are not as active as the other non-mammalian preparations, they still are capable of stimulating the fish thyroid, and bullfrog LH exhibits a slight, but measurable, heterothyrotropic activity (bullfrog LH was less than 4”/:, as active as bullfrog TSH and NIH TSH in a bullfrog TSH bioassay versus 2&259(, as active in Gi//ichthp). This heterothyrotropic activity of bullfrog LH relative to its homologous thyrotropin is weak in comparison with that exhibited by the ostrich gonadotropins or sea turtle LH. These results are surprising since bullfrog LH is the only hormone among those tested which has been demonstrated to possess heterothyrotropic activity in tetrapod species (MacKenzie et al., 1978; MacKenzie. 1980). This provides further evidence for the unique nature of the response of the fish to exogenous hormones. Although the known LH contamination of the bullfrog thyrotropin fraction makes it difficult to draw conclusions about the ability of bullfrog thyrotropin to stimulate the teleost thyroid, an intrinsic activity for this thyrotropin is likely since bullfrog LH alone is much less potent. The low activity of the bullfrog preparations may be due in part to the fact that they are relatively old preparations (approximately three years older than the others) and they have deteriorated with age. It is clear, however. that all gonadotropins tested here possess intrinsic teleost thyroid stimulating activity. The present study expands upon the previous work of Fontaine and others using mammalian hormones. and confirms that heterothyrotropins exist in all vertebrate classes. In addition, these results demonstrate that the specificity of the teleost thyroid response to heterologous pituitary glycoprotein hormones is indeed unique in comparison with that of all other vertebrates tested (Brown & Munro, 1967; Fontaine. 1969: Breneman, 1973; Jackson & Sage, 1973; MacKenzie, 1980). Recent studies on the structure and function of these hormones have shown that although these homologous molecules have undergone considerable evolutionary divergence in function, their structural evolution appears to have been quite conservative (Licht er al.. 1977; Farmer & Papkoff, 1979; Fontaine, 1980). The teleost thyroid receptor, instead of distinguishing these hormones as thyrotropins and gonadotropins. reacts to some common property of these molecules and recognizes them all as thyrotropins. The exact characteristics of these molecules which endow them with teleost thyroidstimulating activity are still unknown. but they appear to be properties of the intact molecule. as subunits are inactive. This teleost response apparently represents a highly derived condition resulting from over two hundred million years of independent teleost evolution (Romer. 1966) and is further evidence that patterns of thyroid specificity have evolved independently within each vertebrate class (MacKenzie rf al.. 1978; MacKenzie. 1980). Although the thyroid gland in a particular vertebrate group may be able to dis-

tlnguish dotropins.

bctuwn

to hetcrolopous Finall).

homolopou~

it is lmpossiblc

and gona-

how It will

react

hormone\.

it should

of thyrotropins

th)l-otrop~ns

to predict

be noted

that

and ponadotropins

lunctional ma!

ovalap

be a more

common

phenomenon than -as once bclIc\cd. Recent work with sensitive assays has shown that the lack 01 thyroid specificity for thyrotropin. first demonstrated in fish with hcterologous hormones. may bc detected in other vcrtcbratcs with both heterologoua and homologous hormones (Amir (‘I (I/.. lY77: MacKenzie c’r trl.. 1978 : Silverbcrg ~‘r (I/.. 1’378: Carag’on 1’1 trl.. IYXO). For this reason, it is interesting to note that tclcost gonadotropins arc the onI4 hormones tested to date which lack heterothyrotroplc actl\ity in tcleosts (Fontaine, 1969: MacKenzie. unpublished results). Their inability to stimulate the teleost thyroid indicates that they may hacc diverged considerably in structure from other vertebrate gonadotropins. and further studies on the structure and receptor interactions of both tcleost gonadotropins and thyrotropins in comparison to tetrapod hormones will be important in elucidating the structural hasls for heterothyrotropic 3ctivit4.

.4~~no~1~/~~dyc,71r,lfs -Hormone preparations were gencrously provided by Dr H. PapkolT. Dr J. Pierce and the Pituitary Hormone Distrihutlon Program of the National Institutes of Health. Thyroxine antihcrum was provided by Dr V. Krusc. National Institute of Animal Science. C‘openhagen. Denmark. Brian McCrecry. Bob Gunther and Richard Nishioka provided valuable assistance with various aspects of these cxperlmcnta I thank Drs Paul Licht and Howard Bern for their \aluablc criticrsm and direction. This work wa\ supported hy NSF Grant PCM-7X-12470 to P. Licht and H. PapkolT REFE;RE\CE;S AMIR S.. U~HI%~L.RA H. & IUGIIAK S. ( 1977) Interactions of bovine thqrotropin and preparations of human chorionic gonadotropin with bovine thyroid membranes. J. c,lirz. Endocr. .Mrtuh. 45. 2X&.292. BR~NE~AI\ W. R. (1973) Bioassay of thyrotropin m chicks with simultaneous cstlmation of gonadotropin. &?I. Camp. Endocri,lol. 20. 41 52. BROWS J. & MUNRO D. S. (lY67) A new III ritro assay for thyroid-stimulating hormone. J. Endwr. 38. 439 444. CARAY~U P.. LI:FORT G. & NISIILA B. (19x0) Interaction of human chorionic gonadotropin and human luteiniring hormone with human thyroid membranes. Endocrinoloyy 106, 1907 1916. CHAN H. H. & EALES J. G. (1976) Influence of bovine TSH on plasma thyroxine levels and thyroid function in brook trout. Salvelinus fonrinalis (MitchIll). Gen. Camp. Endocrrnol. 28, 461472. CCIOPRA I. J. (1978) RadIoimmunoassay of lodothyronincs. In Rudiui,nmuno~r.\.\~~~ (Edited bq AHRAHA~I (3.). pp. 67Y- 703. Dekker. New York. FAKMFR S. W. & PAPKOFF H. (1979) Comparative biochemistry of pituitary growth hormone, prolactin and the glycoprotein hormones. In fformorw\ md Erv~lurio~~ (Edited by BARRINGTON E. J W.), Vol. 2, pp. 525-559. Academic Press, London

dans le determlmsme de la spt:clficlte roologque d’act1on. C.r I,chd Shl,lC~ 41 II‘/. SCI Purrs Serve 111. 292. SO7 5 IO. GRAM, E. G. & STLTSONM. H. (lY77) I‘hyroldal responses to cxogcnous mammallan hormone\ an Flr~ldlti~~\ i~cqc,roc/rru\ (abstracts. .~I,I. %00/ 17, X57 GKALI E. G. & STl.lSON M. H llY7X) Annual rhythm of thyroid sonaltlvlty to TSH 1n the euryhaline teleost, FI~JIdfr/~r.\ i~cf<,roc,/ir~c\ (absrract). In Ccmpwotiw Emlocrirlc~/mq\~(Fditcd by GAILL.ARI) P. J. and BOI R H H.). p. 171 Elsevler. Holland. GRAI. E. G. & STFTS~N M H. (1979) Growth hormone IS thyrotropic in Fttr~t/~rlrr\ hrrtwc/iru\. GUI. Camp Emfoc~r. 39. I x. JA( 6so2i R. G. & %(,I, M I lY73) .A comparison of the ctl’ects of mammalian TSH on the thyroid glands of the teleost Gcdci( /I~/I~..\ /(,/I,\ and the elasmobranch DLI.\\.Q~I\ \&I)I<~. C‘or,f/‘. B/oc,hr,n~. /‘it I WI. 44A, X67 X70. t>lC ,,T P.. PAI’KOFF H.. b’.AK\1lK s. b’. ML~t.1I.R C. H.. ‘rWl CI. W. 6i C-RI b’s D. (lY77) E>o)utlon of gonadotropm structure and function R