Behavioural correlates of genetically enhanced susceptibility to stomach lesion development

J

Psychosomatx

Research,

1961. Vol

5, pp

120 to 126

Pergamon

Press Ltd

Punted

m Northern

Ireland

BEHAVIOURAL CORRELATES OF GENETICALLY ENHANCED SUSCEPTlBILITY TO STOMACH LESION DEVELOPMENT* SINES?

J. 0 (Receded

13 June 1960)

IN AN earlier paper (SINES, 1959) data were presented regarding the Increased susceptlbrhty to stress induced stomach lesions m the rat followmg selective breedmg for two generations In that earlier study rt was noted that ammals which developed stomach lesions appeared to squeal more upon handling, and to resrst handlmg more vlgorously than stock animals Such findings suggested that selection pressure relative to the particular stress response of stomach lesion development might result also m drscrlmmable behavloural changes which could be measured by means of standard observational techniques The present report deals wnh ObJective behavloural data and lesion incidence among stock Sprague-Dawley albmo rats and the S&S,, and S, generations of ammals selectively bred for stomach lesion development METHOD Subjects SubJects (Ss) m this study were 145 rats all of which were 80-90 days of age when subjected to the stress experience The groups of ammals used are indicated m Table 1 All animals were born and raised m this laboratory and had been maintained on standard laboratory regnne whrch involved ad /lbfood and water, mdlvldual housing for pregnant and nursing females, group housing after weamng at 21 days of age, laboratory temperature controlled at 74 f 2”F, and a 12 hr hght-dark cycle TABLE

1 -NUMBER

OF ANIMALS

GROUPS

Group SpragueDawley s, s, &

USED

IN EACH

OF THE

IN THIS Sl UDY

N Males

N Females

20 26 17

22 31

N Total

I ~ ~ ,

42 57

9 72

73

/

~

14s

* This study was supported III part by grant A-2765 (Rl ) M H from the Natmnal Umted States Pubhc Health Servxe t From the Washmgton Unwerslty School of MedIcme 120

Institutes

of Health,

Behavloural correlates of genetically enhanced susceptlbllrty to stomach lesion development

121

Apparatus and procedure

Between 80 and 90 days of age all Ss were observed mdrvrdually for a 2 mm perrod m 5 x 3 feet open field, the floor of whrch was marked off mto 1 x 1 foot squares Behavrour noted m the field consisted of number of squares entered, number of times each ammal reared on Its hmd legs and number of boluses deposrted The first two measures, number of squares entered and amount of rearing are used as mdtcators of actrvrty and the number of boluses JS conventronally employed as a measure of emotronahty m the rat Wnhm 12 hr of the open field experience, all animals were mdrvrdually placed for 12 hr m a wne mesh enclosure which effectively rmmobrhzed the animal lmmobrhzatron for 12 hr was employed wrth subjects on whrch the present report JS based while prevrously 48 or 24 hr of rmmobrhzatron m a plaster of parrs cocoon had been used Forty eight and twenty four hours of plaster cast rmmobrhzatron were found to produce stomach lessons m 100 per cent of S, animals, necessrtatmg the reduction m the severity of the imposed stress m order to allow for contmued selectron S, ammals were injected with an amount of an antrcholmergrc compound sufficrent to prevent lesion development m stock animals rmmobrhzed for 12 hr (SINES, 1960) Followmg the 12 hr perrod of rmmobrhzatron animals were anesthetized with ether and the stomachs inspected for the presence of lesions Stomachs were rated O-4 dependmg upon lesion seventy, with “0” mdtcatmg a normal stomach, “1” “2” nregularrty of the mucosal lmmg, “3” a clearly defined defect small petechrae, and “4” the most severe lesion observed, bemg a which usually mvolved bleeding, palpable defect whrch almost mvarrably mcluded consrderable congealed blood m the base of the crater Selec tron procedure

Three females and two males of the S, group showing severe bleeding lessons were saved for breeding and were the parents of the S, subjects One S, male and two S, females were saved for breeding purposes Since all S, animals showed lessons, the breedmg stock from that generation was selected on the basis of lesion severity Those S4 ammals which were saved had lesson with ratings of “4” The one S, male and the four S, females which were saved for breedmg were those with lessons wrth ratings of “4” S, animals have not yet been subjected to stress RESULTS Stomach lesions

As noted m several previous reports (SAWREY, 1957, SINES, 1959) females m all of the groups showed hrgher mcrdence or severity of stress-induced stomach lessons The difference between stomach lesion mcrdence m stock males and females was srgmficant beyond the 0 01 level, although the differences between males and females m the other groups did not reach a srgmficant level In the S, generation, where mcrdence was 100 per cent m both males and females, the seuerzty of stomach lessons was greater for females, although the difference m severrty drd not reach statrstrcal srgmficance. From Table 2 it can be seen that the mcrdence of stress Induced stomach lessons continued to increase through the S, generation. The data relative to lesson mcrdence m the S, animals are not comparable to the S, and S, data because prior to Immobrlrzatron the S5 rats had been given an anttchohnergrc drug m order to reduce the number 4-mPP)

122

J 0 TABLE 2 -STOMACH

SINES

LESION INCIDENCE IN FOUR GROUPS OF ANIMALS

I

Females

Males

I Group

Sex Leston mctdence

Leston mcrdence

P*

P*

j

~ drfference

!

Srgmficance of sex drfference

I SpragueDawley S, S4 S,

I1

I 20%


71% 100% 25%

45%

1

82% 100%

I

45%

I

to 01
* Stgmficance of comparison with Sprague-Dawley t Stgmficance of compartson wtth Sprague-Dawley ammals thme (regtstercd brand name) where leston mctdence was 0 %

I

+25%

’


i-11%

)

NS NS NS

0%

+20%

premedmated

wtth 0 17 mg/kg

of Proban-

and severrty of lesions Stock ammals so treated with an antrcholemergtc before 12 hr of rmmobrlrzatton do not develop stomach lessons. These data add further support to the orlgmal data on which was based the conclusion that there 1s a srgmficant genetic factor determmmg an orgamsm’s predisposrtion to develop acute ulcerative stomach lesions followmg rmmoblhzatron induced stress. Wergh t

The data found here concernmg body wetght agree, m general, with the trends observed m other inbred strains m that the average body weights m the S,, S,, and S, generations of lesson susceptrble ammals (both males and females) were srgmficantly Of particular interest smaller than the origmal stock from whtch they were drawn however, was the tendency, first seen m the S, females and clearly present m both males and females of the S, generation, for werght to regress toward the mean werght of the stock animals (see Table 3) In another study m this laboratory of several pure strains of the laboratory rat, the rank order coefficient of correlatton between mean weight and number of generaIn view of such data the tions of brother by srster matmg was found to be -0.90 postulatron of any relatronshrp between weight and susceptrbrhty to stomach lesion development seems unwarranted at this pomt. TABLE ~-COMPARISON

OF

MEAN

BODY WEIGHT(IN

Males ~~

~RAMS)OF FOUR GROUPSOF

Females _ ~~~

Werght

SpragueDawley S, S, S, * Stgmficance

I ,

Group

324 6 2684 259 2 268 9 of compartson

p*

Wetght

I i

ANIMALS

~

Sex dtfference

Significance of sex dtfference

I

-

’

203 1

;

98 6 82 3 779

wtth Sprague-Dawley


Behavloural correlates of genetically enhanced susceptlblhty

to stomach lesion development

123

The differences between mean weights of the males and females were statrstrcally hrghly srgmficant m each group although the magnitude of the difference decreased regularly. Openjield

behavlour (summarized

m Table 4)

Number of squares entered. Although S, and S, males entered a srgmficantly greater average number of squares m the open field than drd stock males, the average number of squares entered by the S, ammals was not statrstrcally drfferent from that

TABLE

4 -COMPARISON

OF SPRAGUE-DAWLEY

AND LESION SUSCEPTIBLE ANIMALS

ON OPEN FIELD BEHAVIOUR

,

Females

Males

Group

I Slgmficance of sex differences

Sex 1dlfferencest

I

, Squares

P*

Squares

<005 to 01 NS

;

28 32 31 24

P*

1

Rearmg

p*

j SpragueDawley S, S, S,

I ~ I I

19 0 25 3 29 1 22 4 Rearmg

SpragueDawley S, S,

I I i

~ 5

~

NS P*

-96 -68 -27 -17

1

’

70 107 13 1


91 I
Defecation

20 47 48 31

P*

(001
104 12 8 11 8 119

/ )

Defecdtlon

1 1 /

06 30 23 31


1

p*

I

to 01
_.

to 05
_.

I

s6 I

SpragueDawley S, S, S,

/

I

6 1 8 1

-34 -2 1 $13 -28


/

to 05

~

+14 117 +25 00

~ ~

NS NS to 05 NS

* Slgmficance of comparison with Sprague-Dawley values t Plus signs Indicate higher mean scores for males

of stock males This regression by the S, males toward the mean number of squares entered by the stock males paralleled the slmrlar but smaller trend noted above m body weight Among females, the average numbers of squares entered by the thn-d, fourth, and fifth generations of lesion susceptible animals were not srgnificantly different from the average of the female stock animals. The average number of squares entered by the S, females was lower than the female stock value (not srgmficant) but rt was srgmficantly lower than the female S, average (0.01 level of significance). Rearmg. The average amount of rearmg in the open field by S, and S, males was

124

J 0

SINES

sigmficantly greater than the average amount of rearmg done by the stock males Here again, however, the regression toward stock levels by the S, animals was noted Although the S, females reared more often m the open field than stock females, (0 05 level), the S, and S, females regressed slightly to levels not significantly different from the mean of the stock females In the present study both of the above measures, number of squares entered and amount of rearing, were considered to be measures of gross motor actlvrty Thus, as can be noted m Table 3, successive selectively bred generations of leston susceptible animals tended to manifest Increased activity when subjected to the novel, mildly stressful open field sltuatton. The tendency for htgher levels of motor activity to characterize those ammals more susceptible to stomach leston developments 1s further supported by the sex differences where rt was found that females tended to manifest both higher activity levels and higher lesion mcrdence Defecation With regard to defecation, as was the general case wrth the actlvrty measures, the thud and fourth generatton susceptible males showed slgmficantly higher mean scores, while the mean for the S, group was not stgmficantly larger than the stock animals mean scores. There was, among all three generations of lesion susceptible females tested, srgmficantly more defecation than the stock animals showed Llttev size and dehery

among lesion susceptible

anrmals

During the two and one half year period m which this research has been under way, stock animals have been bred and ratsed m this laboratory m order to ensure a populatton of stock animals with pre-stress experimental backgrounds as similar as possible to those of the selectively bred animals Although records have not been kept systematically, the average litter size for stock animals has been between 8 and 12 pups, with the range of from 6 to 17. None of the stock females have died m labor or m delivery The average litter size m the S,, S, and S, generations has tended to be smaller, 5 7, 5 0,7*0 respectively Whether the differences between stock and the experimental animals m mean Inter size are statlstlcally significant cannot be determined at this point. The trend noted, however, suggests that there are other basic characterrstlcs also m which stock animals may differ from animals which are susceptible to stress mduced stomach lesions The frequency with which females of the selectively bred generations have died m labor or during the delivery of then pups has been striking Two of the S, females died while dehvermg then third litters, at approximately 280 days of age One S, female died dehvermg her second litter at approximately 210 days of age and a second S, female died during the delivery of her thud litter when she was about 280 days of age At the age of 280 days the stock females m this laboratory have been able to deliver litters of average size with no apparent difficulty Neither of the two S, females appeared to have serious trouble m delivery but to date, two of four S, females have died during delivery. Another observation mvolves the tendency for lesion-susceptible females, which do deliver then pups, to ignore then young The causes for such non-maternal behavror are not clear at this time but may have specific srgmficance m view of some of Cannon’s work (quoted m BEST and TAYLOR, 1945, p 945) Cannon found that

Behawoural

correlates of genetlcally enhanced susceptlblhty

to stomach lesion development

125

“ktttens deprived of then- sympathetic trunks grow normally, female cats become pregnant and gave birth to young, though the mammary glands do not functzon and the maternal mstmct 1s Iackmg” (present authors’ Italics). It may be that the selection pressure here employed has resulted m animals which are, to some degree, functtonally “sympathectonuzed”. Such IS our present working hypotheses developed to accommodate the data reported here, Cannon’s findings, and data we have reported elsewhere (SINES, 1960) relattve to drug effects on lesion mctdence m rats Fmally, one of the S, females was m labor for over 8 hr with her second litter and appeared close to death when her pups were delivered surgically. She was found to have 8 fetuses, only 4 of them fully developed On the basts of these addtttonal unsystematic observattons, further studies of the pregnancies and deliveries wtll be made DISCUSSION

The data presented here lend support to a prehmmary observation that rats which develop stomach lesions followmg the stress of rmmobrhzatton are more active and emotronal Further, successtve generations of animals selectively bred for lesion development were srgmficantly smaller m body werght than the stock from which they were ortgmally drawn The tendency for the S, and S5 generations to regress toward the stock ammals m almost all of the above characterrsttcs may be the result of the selection procedures employed, parttcularly with S, arnmals The fact that 100 per cent of the S, ammals demonstrated stomach lesions made rt necessary to select breeding stock from that group on the basts of lesion seventy, a criterion which was without doubt less dtscrrmmatmg than an all-or-none crrterron For that reason the use of anttcholmergtcs to retard lesion development among the susceptible animals was begun wtth the S, animals Theassumptron was made that only highly susceptible animals would develop stomach lesions under stress rf an appropriate amount of a drug which IS specrfic for the control of stomach lesions has been used. Data supportmg the use of anttchohnergtcs to control stress induced stomach lesions n-r the rat have been presented elsewhere (SINES, 1960) The strrkmg tendency for lesson suscepttble females to cast smaller litters and to have an unusually high fatality rate during delivery was unexpected and IS to be studied closely m future generations Attentton 1s also presently being directed toward the study of other characterrsttcs such as the probably drfferences m phystologrcal response patterns and possible differences in learnmg abthty between lesion susceptible and stock animals, characteristics m which there may also be changes as a result of the sort of selectton pressure used On the basis of the data presented above one may state that ammals, of this specres at least, which develop stomach lesions when exposed to a gtven amount of stress doffer from others which do not develop stomach lessons when subjected to obJectively identical amounts of stress m that the former animals (1) show more spontaneous exploratory acttvtty m a novel sttuatton, (number of squares entered and amount of rearing m the open field) (2) are more emottonal m response to a novel situation (defecation m the open field) (3) tend to produce smaller Inters

126

J 0

SINES

(4) are offsprmg of females which both* (a) have greater drfficulty dehvermg pups and (b) fall more often to mother the pups which are born.

live

SUMMARY

Data are presented which Indicate that the S,, S,, and S, generations of rats, selectively bred for susceptrbrhty to stress-induced stomach lesions, are srgmficantly lighter in body weight as well as more active and more “emotronal” m an open field situation than are randomly selected Sprague-Dawley albmos The selection procedures employed have continued to result m greater incidence of stress induced stomach lesions although a tendency was noted for the S, and S, animals to regress toward the levels of stock animals m some of the behavroural characterrstrcs studied There appears to be a tendency for the stomach-lesion susceptible females to cast smaller litters and therefore to be an unusually high proportion of lesion susceptible females to dre during dehvery The tendency noted for lesson susceptrble females to ignore then pups may be related to Cannon’s earlier work on sympathectomrzed animals. A number of leads which require continued study were mentioned REFERENCES

BESTH C and TAYLORN B (1954) Thephyslologrcal basis of medlcalpractlce (Fourth Ed ), p 945. Wdhams and Wdkms, Baltimore SAWREYW L , CONGERJ J and TURRELLE S (1956) An experImenta mvestlgatlon of the role of psychologlcal factors m the productlon of gastric ulcers m rats J camp Physlol Psychol, 49,457. SINES J 0 (1959) Selective breedmg for development of stomach lesions followmg stress m the rat. J camp Phynol Psycho1 52, 615 SINES J 0 (1960) Experlmental productlon and control of stomach lesions m the rat J Psychosorn. Res 4. 297