activity the pars interinedia and pars nervosa of the fresh ox pituitary collected material, dried and powdered in a mortar, is used as a standard

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1 THE PHYSIOLOGICAL ACTIVITY OF THE PARS INTERMEDIA AND PARS NERVOSA OF THE OX PITUITARY QUANTITA- TIVELY COMPARED. By P. T. HERRING. (From the Physiology Department, University of St Andrews.) (With six figures in the text.) (Received for publication 29th April 1914.) WITH the object of making a quantitative analysis of their physiological activity the pars interinedia and pars nervosa of the fresh ox pituitary have been isolated in the nmanner described in a previous paper.' The collected material, dried and powdered in a mortar, is used as a standard for making up extracts of different strengths in Ringer's solution. It may be objected that the dried gland varies in activity, and does not constitute a trustworthy standard. In my experience the dried material exhibits remarkably constant results when made up into extracts of the same strength, and loses little, if any, of its activity after being kept for months in the dried form. For the purpose of comnparing the activity of the pars intermedia and pars nervosa quantitatively, the two parts have been removed from the same pituitaries, and subsequently treated in exactly the same manner, so that I think it may be conceded that extracts of the two may be fairly compared in their results. The action of extracts of the two portions has first been noted upon the isolated uterus of the virgin rat, and thereafter upon the blood-pressure, kidney volume, and urinary section of aniesthetised cats. ACTION OF EXTRACTS UPON THE ISOLATED UTERUS. The uterus of the virgin rat has been used in these experiments in preference to that of the guinea-pig. I have found the uterus of the guinea-pig to be too sensitive, and apt to go into a state of tonic contraction in Ringer alone. This is probably due to the fact that the guinea-pigs were too old, and not virgin. Dale and Laidlaw 2 state that their best results were obtained from virgin guinea-pigs of not more than 350 gm. weight. I have been unable to procure suitable animals, and, moreover, the uterus of the guinea-pig is rather large for the testing of small quantities of extracts. The virgin rat of from three to four months old furnishes a small uterus which has little rhythm of its own in Ringer, and gives very constant responses to repeated immersions in the same strength of extract. I P. T. Herring, Quart. Journ. Exper. Physiol., This volume, p Dale and Laidlaw, Journ. Pharm. and Exper. Therap., 1912, iv. pp. 75 to 95.

2 268 Herning The uterus was suspended in oxygenated Ringer at 380 C. in the manner described in the previous paper (op. cit.); with this difference, that both horns of the uterus were used instead of only one. The lever was soc arranged as to magnify the extent of contraction by two, and was suitably weighted by the aid of a few preliminary experiments. Various strengths of extracts were employed, and it was soon apparent that weak extracts give results which are constant and comparable. Strongextracts provoke a tonic contraction which is only slowly relaxed during the subsequent immersion in fresh oxygenated Ringer. A number of important details have to be attended to, which are enmphasised in Dale and Laidlaw's paper. One of the most important is the constant renewal of the Ringer's solution, which, if neglected, results in increased sensitiveness of the uterus, and a consequently bigger contraction to the same strength of extract. Both the Ringer's fluid and extracts were oxygenated by a constant stream of 029 and their temperature kept at 380 C. The changes of Ringer were raised to that temperature only immediately before use. The uterus was immersed for 15 seconds at a time in the extracts, and well rinsed in fresh Ringer after each contraction. A *01 per cent. extract of the dried pars intermedia was taken as the standard with which to compare extracts of the pars nervosa. This strength was selected after a nuinber of experiments in which stronger solutions had been employed. A -01 per cent. extract of pars nervosa was then diluted with Ringer into a series of extracts of weaker percentages, and their action upon the uterus compared with that of the *01 per cent. extract of pars intermedia. Fig. 1 shows the action upon the rat's uterus of-(a) a *01 per cent. extract of pars intermedia; (b) a 005 per cent. extract of pars nervosa; (c) a *002 per cent. extract of pars nervosa; (d) a *00125 per cent. extract of pars nervosa; (e) a *001 per cent. extract of pars nervosa; (f) a 0005 per cent. extract of pars nervosa. Taking the *01 per cent. extract of pars intermedia as a standard, the extract of pars nervosa employed in (b) is j that strength; in (c).1th; in (d) Ith; in (e) IJ th, and in (f ) -1J th. All the contractions recorded in fig. 1 are by the same uterus, but were not taken in the order given. The uterus responded uniformly to the same extract, and the height of the contraction varied directly with the strength of the extract employed. A series of observations with other uteri, and with other extracts similarly made up, gave the same results. The contraction produced by an extract of pars intermedia was reproduced in height by extracts of pars nervosa of from a half to a fifth of that strength. It appears, therefore, that in its action upon the uterus the pars nervosa is from two to five times as strong as the pars intermedia. It is certainly not ten times as strong. In fig. 1, (e) is a distinctly smaller contraction than (a). The remarkable sensitiveness of the uterus to extracts of the posterior lobe of the pituitary body is clearly evident. A.0005 per cent. extract of dried pars nervosa produces a very distinct response (fig. 1 (f)), and even considerably weaker extracts are active.

3 Activity of Parts of Ox Pituitary 269 Strong extracts of both pars intermedia and pars nervosa stimulate the uterus to prolonged contraction. The actual height of the contraction may?t d J FIG. 1.-Effect of various strengths of extracts of the pituitary body of the ox upon the isolated uterus of a virgin rat. Time of immersion in each tracing is 15 seconds. The upper tracing, ts, represents the uterus; J, the signal; t, the time in 30-second intervals. The excursion of the lever is x 2. a, effect of immersion in a.01 per cent extract of dried pars intermedia in singer * b, effect of a 006 per cent. extract of dried pars nervosa; c, effect of a *002 per cent. extract of dried pars nervosa; d, effect of a *00125 per cent. extract of dried pars nervosa ; e, effect of a *001 per cent. extract of dried pars nervosa; f, effect of a *0005 per cent. extract of dried pars nervoss, be no greater than that produced by weaker solutions, but the uterus remains tonically contracted for a longer period. In this respect extracts of pars nervosa appear to have the more pronounced action. It is sometimes so marked as to be suggestive of the presence in the pars nervosa of some additional active principle.

4 270 Herring cr 4. c4d._2 n Ca ce --' cli

5 Activity of Parts of Ox Pituitary.271 ACTION OF EXTRACTS UPON THE BLOOD-PRESSURE, THE VOLUME OF THE KIDNEY, AND THE SECRETION OF URINE. The animals experimented upon were cats, which were anesthetised with chloroform, and received in addition an intravenous injection of chloral hydrate. The blood-pressure was recorded from the right carotid artery; the left kidney was placed in an oncometer connected with a piston recorder; and the flow of urine was registered by a tube in the bladder conveying the drops to an electrical marker. It has already been shown (op. cit.) that in the cat, at any rate, extracts of pars intermedia of a strength equivalent to 2 per cent. of the fresh gland a k FIG. 3.-Effect of an injection into the jugular vein of a cat of 2 c.c. of a *5 per cent. extract in Ringer of dried pars intermedia of the ox pituitary previously treated with absolute alcohol. have no specific effect upon blood-pressure, kidney volume, and urinary secretion. Repeated experiments confirm this observation. Fig. 2 records the action of a first dose of 3 c.c. of a 1 per cent. extract of dried pars intermedia, the identical extract which, after being diluted ten times with Ringer, was used to produce the contraction of the rat's uterus in fig. 1 (a). After a slight preliminary fall of blood-pressure there is a slight rise, which is continued for some time. The kidney shows little alteration, and the flow of urine is, if anything, diminished. The same slight rise of bloodpressure and dilatation of kidney were in this cat brought about by the injection of 3 c.c. Ringer alone. The action of a 5 per cent extract of pars intermedia is shown in fig. 3. This extract is one from which the alcohol-soluble material had been

6 2d72 Herriiio, removed b)v repeated extractioni of the dried gland with absolute alcohol. The inijectionl of 2 cc. has In action upan blood-pressure. Trlhe temporary Fi(;. 4. -Effect of an ilijectioll ilito the j ngular veini of the samne cat of 2 c. c. of a *01 per cent. extract in Ringer of (dried pars nervosa of the ox p)ituitary. I'Tiis tr-acing is a continuation of tiff. 3, Fin(. -Elrect of an inijectionl ilit( the jntgulair vein of a cat of 3 c. c. of a *01 per cei't. extitact in Riniger of (trie.d pars nervosa of the ox pitnitary. Tue zein> of b)lood-pressnre corresponds witl the line t. expansion of the kidney is caused by the injection, but is Inot specific, and there is no increase in the secretion of uriine-if aniythinig, the flow is slightly

7 Activity of Parts of Ox Pituitary.273 decreased. Eight minutes after this injection a dose of 2 c.c. of a *01 per cent. extract of pars nervosa was given. The effect is shown in fig. 4, which is a direct continuation of fig. 3. There is a rise of blood-pressure, marked expansion of the kidney, and distinct increase in the flow of urine. Extracts of dried pars nervosa of a strength of *01 per cent. produce the typical effects of extracts of the posterior lobe of the pituitary body upon the blood-pressure, kidney volume, and urinary secretion of cats. Fig. 5 shows the results of the injection of 3 c.c. of a *01 per cent. extract of pars nervosa in a cat with a low blood-pressure. The expansion of the kidney is particularly marked. No urine was being secreted before the injection, but a flow is produced by the action of the extract. a _i FIG. 6.-Effect of ail injection into the jugular vein of a cat of 3 c.c. of a extract in Ringer of dried pars nervosa of the ox pituitary. 005 per cent. Similar, though less marked, results are given by the injection of 3 c.c. of a 005 per cent. extract of the dried pars nervosa (fig. 6). Extracts of a weaker strength, e.g. *001 per cent., injected in doses of 2 to 3 c.c., have a little action, but cannot be depended upon to produce a typical effect. From the above-recorded experiments it is evident that the kidney is extreinely susceptible to the action of extracts of pars nervosa, a fact which was stated by Schafer and Herring in The kidney reaction is not obtained by such dilute solutions as suffice to stimulate the isolated uterus, but the mode of application is quite different. In the case of the uterus the organ comes into direct contact with a quantity of fluid of a definite strength. In the case of the kidney 2 or 3 c.c. of the extract are diluted by the blood of the animal, and it is impossible to say what is the strength of the extract by the timne it produces its effects; it must, in any case, be much less than that of the injected fluid.

8 274 Activity of Parts of Ox Pituitary SUMMARY. Both pars intermedia and pars nervosa of the ox pituitary contain an active principle which stimulates uterine muscle, but extracts of the two are not equally active. The pars nervosa is from two to five times more powerful than the pars intermedia in its action. The pars intermedia, in extracts of *5 per cent. and under of the dried material, has no specific action upon blood-pressure, kidney volume, or urinary secretion. On the other hand, extracts of pars nervosa of a strength of 005 per cent. of the dried material produce, in doses of 2 to 3 c.c. iinjected intravenously, the prolonged rise of blood-pressure, increase of kidney volume and of urine, characteristic of extracts of the posterior lobe of the pituitary. If the active principles producing uterine contraction, and the specific action upon blood-pressure and kidney, are one and the same substance, one would expect that an extract of pars interinedia of from two to five times the strength of an extract of pars nervosa would have a similar action upon blood-pressure and kidney as it has upon the isolated uterus. But this is not the case. An extract of pars intermedia of a hundred times the strength of that of the pars nervosa fails to give the typical action of extracts of pars nervosa upon the blood-pressure and kidney. Dale and Laidlaw (op. cit.) state that they have never found an extract which was inferior in pressor action and had not a like inferiority as a diuretic and a uterine stimulant. This may be true of extracts of the pars nervosa, but does not hold in a comparison of the extracts of pars intermedia and pars nervosa. The evidence is not sufficient to prove that there are two separate active principles in the posterior lobe of the pituitary body, but it is at least suggestive of that being tbeftase. The probability is that the active material of the posterior lobe of the pituitary is a product of the epithelial cells of the pars intermedia, and that there are stages in the breaking down of these cells in the pars nervosa. The substance acting upon the uterus is formed at an early stage in the cells of the pars intermedia, but the substance acting uponi blood-pressure and kidney is a later product resulting from the breaking down of the hyaline bodies or disintegrating pars intermedia cells in the pars nervosa. The expenses of this research have been defrayed by a grant from the Carnegie ftrust.