The Discovery of Serotonin Irvine H. Page Perspectives in Biology and Medicine, Volume 20, Number 1, Autumn 1976, pp. 1-8 (Article) Published by Johns Hopkins University Press DOI: https://doi.org/10.1353/pbm.1976.0058

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PERSPECTIVES IN BIOLOGY AND MEDICINE Volume 20 · Number 1 · Autumn 1976

THE DISCOVERY OF SEROTONIN IRVINE H. PAGE, M.D*

It may well be asked, What difference does it make in the long run who discovered what and when? My answer reflects my own deep biases and experiences with particular discoveries, in this case, serotonin. I believe that science is best taught and remembered when it is humanized and thereby related to the people involved. This is simply good pedagogy, part fact and often part poetic license. How much of each history

will judge. A firsthand, eyewitness report could provide the substance for debate. This has always been the way of science, and I hope it always

will be. But there are other reasons ás well.

It has now been 28 years since serotonin was isolated and synthesized

and nearly 40 years since the presence of a uterine-contracting substance was found in extracts of gastrointestinal tract. In the long interim, it appears that, "after all is said and done," there should be no need to say

more! But I am doing so, because the record itself is clear; it has simply not been carefully read or has been distorted. Let me amplify. As most investigators should know but often fail to show that they do, to describe certain properties of a mixture of substances or even to "purify" is not

synonymous with "isolating," "identifying," and determining the chemical structure of a substance. Thus many bioactive substances have been described in terms of certain actions but have not been isolated, al-

though they have been concentrated. Often such a procedure can be just as useful as actually identifying the material. Insulin was used long before

its true chemical nature was known. But there have also been endless

extracts of organs containing all manner of substances which have led only to further confusion, since their actions were only the sum of sev-

eral active agents. This was at the heart of the problem of serotonin.

*Research Division, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland, Ohio

44106.

Perspectives in Biology and Medicine · Autumn 1976 | 1

What Serotonin Has Meant to Me

In 1928 at the close of my hospital internship, an unexpected offer was. made to me by Geheimrat Richard Willstätter, adviser to the Deutsche Forschungsanstalt für Psychiatrie in Munich. He asked if I would form a chemical division in that institute to study the "chemistry

of the brain." I had graduated in chemistry from Cornell University in 1921 and had been provided with what I thought to be a first-rate education by very distinguished chemists, for which I shall be ever grate-

ful. But to initiate a discipline of which I had never heard and soon found that few others had, either, was a venture only the young and

inexperienced would undertake. That is why I did it! Three years in Munich was marvelously stimulating, with such intellectual giants as Wieland, Fischer, Willstätter, and Fajans, to mention but a few. Then there was the more casual association with the great at the

headquarters of the Kaiser Wilhelm Gesellschaft in Berlin-Dahlem. Such a panoply during this magnificent period of German science needs only the mention of Einstein, Schroedinger, Warburg, Plank, and Haber to underscore its greatness.

Another aspect of this period was the opportunity, since travel was easy between Germany and England, to know men of renown such as Dale, Dudley, Gaddum, von Euler, Verney, Evans, Drummond, Young, Sherrington, and Florey. These were true scientists, uncontaminated by

administrative and financial distractions which have subsequently so weakened the fiber of our "greats."

With all this name dropping, it might be concluded that, as director of the Chemical Division, I too might share their nimbus. Time and the printed record, alas, show that my approach was sound but pedestrian. Only two things were accomplished: "brain chemistry," or neurochemistry, with the help of men such as Himwich, Elliott, and Quastel, began to

appear dimly as a reality; and my book, The Chemistry ofthe Brain, was written, which was none too soon, for the first one ever written on the

subject was by J. L. W. Thudichum in 1884. This was all to have great personal importance to me later in our work on serotonin. When I emigrated to the Rockefeller Institute Hospital to work on nephritis under Dr. Donald D. Van Slyke, a wholly new panorama opened before me. My acquaintance with cardiovascular disease had

begun with the preparation of a student thesis under the watchful eye of Dr. James Ewing on the histopathology of blood-vessel disease, as seen in the large, municipal Bellevue Hospital. Dr. Ewing was our beloved teacher of pathology at the Cornell Medical School. So at Rockefeller Hospital my interest turned again to cardiovascular

disease. But that is another story. What is pertinent is that, in undertaking the study of the hypertension of nephritic patients, my future course 2 I Irvine H. Page · Discovery of Serotonin

was unconsciously being set. Being at heart a chemist, I instinctively looked for a humoral agent responsible for the elevated blood pressure. From 1931 on, the search continued without interruption. Through playing tennis, I made a fortuitous contact with the very gifted Robert Elderfield, an outstanding figure in the field of indolalkylamines. I studied many samples of these substances in connection with their effects on the cardiovascular system. So strange were some of their names that theJournal ofPharmacology and Experimental Therapeutics published little of this work, doubting, I suspect, that such substances actually existed. Arthur Corcoran and I moved to Indianapolis City Hospital (Lilly

Laboratory for Clinical Investigation) in 1937 and continued the search for pressor and depressor substances, in association with Oscar Helmer

and Kenneth Kohlstaedt. It had been apparent for about a century that a very disturbing phenomenon occurred when blood clotted: one or several vasoconstrictor substances appeared. This led early physiologists to use for perfusion experiments blood that had been defibrinated and passed through a pair of lungs to remove the constrictors. But first let us turn back nearly a century to observe the slow growth of thinking and research on this subject.

It was in 1868 that Ludwig and Schmidt [1] noted that defibrinated blood increased vascular resistance in perfused dog muscle. Prevention of clotting with sodium citrate abolished the vasoconstrictor action of serum. Janeway and Park [2, 3] in 1912 and Stewart and Zucker [4] in 1913 showed clearly that ox carotid-arterial strips contracted only after

blood had clotted. But, from this period on, a number of papers appeared, most of them contradictory of each other. Some of the confu-

sion seemed to abate when Dittler [5] in 1918 thought he could demonstrate two smooth-muscle agonists in serum by dialysis, one stimulating and the other inhibiting rabbit intestine. Hirudinized blood prevented

their occurrence. Doubts were raised because the stimulating substance was claimed to come from red cells. This was partially corrected by Hirose's demonstration [6] that a vasoconstrictor action appeared in

amounts roughly proportional to the platelet count. But, again, confusion appeared because Freund [7] in 1920 found the agonist in plateletfree plasma! The problem became of more interest but further confused when Bayliss and Ogden [8] in 1932 called attention to the "vasotonins" as

substances to be removed in pump-oxygenator-kidney preparations. This led to the general use by physiologists of interposition of a pair of lungs to prevent development of vasoactive materials. Zipf and Hülsmeyer [9] in 1933 described toxic materials formed in blood on

standing which produced hypotension and intestinal inhibition that was attributed to adenylic acid and another substance that stimulated intes-

Perspectives in Biology and Medicine · Autumn 1976 | 3

tine and uterus but was vasoinactive. Freund [10] identified the latter as

tyramine. The chaos was all but complete when Simon [11] claimed that the agonist was not the same in different animal species! Two now forgotten pharmacological observations were made: one, by Heymans, Bouckaert, and Moraes [12] in 1932, was that ergotamine reversed the vasoconstrictor action of what must have been incompletely

defibrinated blood; the other, by Bing [13], was that large doses of ergotoxin injected into a kidney-perfusion circuit elicited a quick decrease, followed by an increase, in renal blood flow lasting several hours.

This, in outline, was the course of thought and investigation concerned primarily with the vascular actions of this strange phenomenon resulting from coagulation of blood. Another line of thought was concerned with the intestinal action of

such substances. The early literature is diffuse and contradictory, as it is

in the case of the vasoactive materials. But more consecutive work began

in 1933, when Vialli and Erspamer [14] gave the name "enteramine" to a substance or substances contained in extracts of rabbit gastric mucosa which took on a color after coupling with the diazonium salt of p-nitroaniline. The material appeared to originate in what they called "enterochromaffin," argentophile cells found in the gastrointestinal tract of mammals. They considered the activity to be involved only with the gut, hence the name "enteramine." Chemically it was believed to be a di- or polyphenolic amine. An inactive form of enteramine was also

found which was activated by heating at pH 7-10. These extracts of gastrointestinal mucosa containing enteramine stimulated intestinal and uterine strips after atropinization and reduced blood pressure in atropinized rabbits and cats. Much of this work over many years appeared in a great variety of journals but, except for the work of the gifted Erspamer, had no continuity. It is not surprising, therefore, that those of us interested in cardiovascular physiology were unaware of what had been done and the little we knew we discounted as being utterly chaotic. It was crystal clear that, if a pressor substance was to be isolated from the blood of hypertensives, the problem of vasoconstrictor materials resulting from overt or even occult coagulation had to be overcome. This was the motivation for our work on serotonin.

A number of false starts were made. I had searched for a pressor

substance, or lack of it, in the urine of my hypertensive patients while at Rockefeller Hospital. After going through all the tedious steps to get rid of salt and still be able to concentrate the extract, I finally found what I then thought to be the key to the problem. The extracts of urine of my hypertensive patients contained no pressor substance, but those of the normotensive controls, namely, other staff doctors, contained large amounts. In short, hypertensives retained their pressor substance, and normal persons did not! The bubble was burst when the pressor material 4 I Irvine H. Page · Discovery of Serotonin

proved on analysis to be nicotine, which heretofore had not been known

to be excreted in the urine. A strict head nurse of the old Johns Hopkins tradition forbade smoking as the work of the devil among the patients but was less persuasive among the doctors, from whom the devil was much harder to exorcise.

Another dry run was the search for a pressor substance in plasma ultrafiltrates claimed to be pressor in patients with hypertensive toxemia.

We could not confirm this claim even when the discoverer worked with me for several months in New York.

I had improved a cheap method used long ago for student demonstrations [15]. Others strongly disagreed about its reliability. The ears of rabbits were perfused with a pulsating pressure through the central artery and the effluent drops counted. When coagulation was not allowed to occur or the platelets were re-

moved, no vasoconstriction in the rabbit ear vessels occurred. But even

dilute serum gave strong vasoconstriction. In a sense, then, the unsophisticated rabbit-ear-vessel preparation was the key to the isolation of a substance that had undreamed-of properties. At the time, the method received erosive criticism. Using blood pressure as a measure probably would not have been successful, because we

now know how capricious the response of the arterial pressure is to serotonin, aside from the large quantities of test substances that would be needed.

When several of us moved from Indianapolis to Cleveland to start a research division at the Cleveland Clinic, the problem came with us. We were joined by a very capable young chemist, Dr. Maurice Rapport, the technically experienced Dr. Arda Green, and the ever constructively

critical Dr. James McCubbin. It was this happy circumstance that allowed the problem to be carried to its final solution [16-19], announced in 1947.

At that time we had never heard of Vialli and Erspamer's paper published in 1933 on extracts of gastric mucosa; and, if we had, it could easily have been misleading, because the pharmacological properties, origin of their extracts, and the suggested chemical nature as a di- or polyphenolic amine were quite different from those needed to solve our problem.

A word should be said at this point about the question of priority, raised not by Erspamer and myself but by others. By now the record

should have been set straight but obviously has not. In retrospect, this confusion resulted from the very human trait of not taking the trouble

to read the original publications and, as I have said, the failure of most people to differentiate between isolation and chemical identification of the structure of the substance on the one hand and, on the other, its

separation by extraction, along with many other substances. To my great surprise, my old friend Gaddum was one of the worst and Perspectives in Biology and Medicine · Autumn 1976 I 5

most influential offenders. He repeatedly made this error and confused the nomenclature by declaring that enteramine and serotonin were identical, describing them as being the same substance long before even any similarity was evident. He selected the name 5-HT, an unfelicitous acronym to say the least. A "5-HTogenic neuron" is a monstrosity! Once the active substance was isolated and crystallized and its struc-

ture determined, it was possible for Hamlin and Fischer [20] at the Upjohn Company to synthesize large quantities of serotonin and make it available to all investigators. Another dimension was thereby added. McCubbin and I published encouragement to pharmacologists under the title "Serotonin, or Tenure for the Pharmacologist" [21].

McCubbin's and my immediate interest centered chiefly on the cardiovascular actions of serotonin, since it was these actions that led to its

isolation because of its interference with the then very unsatisfactory

methods of measuring angiotensin.

We found the vascular action of serotonin so variable that we did the

inexcusable—we coined a word, "amphibaric," to describe its multiple effects on blood pressure. These could only be described as bizarre, since, when the neurogenic tone of the test animal was high, as in neurogenic hypertension (debuffering), serotonin lowered blood pressure sharply; and contrariwise, when tone was low, as after destruction of part of the spinal cord, it raised it [22].

McCubbin, Kaneko, and I [23] followed up the observation that the probable mechanism controlling reactivity to serotonin is the degree of

activity of the autonomic nervous system. It was shown that stimulation

of the lumbar sympathetic trunk caused vasoconstriction of both the large and small arteries of a dog's hind limb. When serotonin was in-

jected during the stimulation, the constricted small arteries relaxed, but the larger ones constricted further; the net effect was decrease in total vascular resistence. In the absence of stimulation, serotonin caused con-

striction of both large and small arteries and increase in vascular resistance. These experiments showed the remarkable interplay of neuronal and humoral vasoactivity. Another vascular aspect of serotonin blossomed because of newly aroused clinical interest in carcinoid tumors. Chiefly due to the work of the gifted team of Sidney Udenfriend, Herbert Weisbach, and Albert

Sjoerdsma, Corcoran and I reported along with them [24] on three patients with malignant carcinoid, demonstrating the huge excretion of 5-hydroxyindole acetic acid, a metabolite of serotonin.

The Relationship to Neurochemistry By 1955 the field began to move so fast that the simple procedure

using perfused rabbit ear vessels began to seem amateurish, in the face of the growing sophistication of research stoked by taxpayer largesse. 6 I Irvine H. Page · Discovery of Serotonin

Adumbrating my fate, I wrote two reviews [25, 26] and a small book [27] and fled. However, I was not to be let off scot-free, because of the

finding of serotonin in the brain and the then mild excitement created

by Albert Hoffman's synthesis of lysergic-acid diethylamide and his recognition of its hallucinogenic properties, as well as Gaddum's simple observation that LSD was a serotonin antagonist on intestine. As I explained, while a chemical division of a psychiatric institute was

formed in Munich in 1928, it failed to galvanize the potentially enormously important field of neurochemistry. It merely gave it a shove.

When serotonin had been isolated and synthesized, it seemed obvious enough to examine systematically the tissues of the body to see if it was

present in structures other than blood platelets. Betty Twarog and I [28]

did this and, to our astonishment, found it in relatively large amounts in the brain. So in 1953 the circle was closed. I had entered the circle in

1928, lost my way, but returned via serotonin to neurochemistry.

With the active participation of Drs. Peter Gessner, Merlin Bumpus, Robert Taborsky, and William Mclsaac, a program was initiated to study the metabolic and behavioral aspects of a variety of serotonin derivatives, some of which had been synthesized in our laboratory [29-31]. While many very interesting observations were made on the multiple actions of these derivatives, we soon realized that we could not keep pace with the experts in the behavioral and neurological sciences. As a spinoff, McCubbin, Kaneko, and I [32] found that the acute cardiovascular

effects of reserpine were mediated by serotonin and/or norepinephrine, since reserpine, serotonin, norepinephrine, 5-hydroxytryptophan and 3,4 dihydroxyphenylalanine had qualitatively the same cardiovascular effects when injected into a lateral ventricle or into the cisterna magna.

Because of this, I felt compelled to write postscripts on these aspects in "Chemistry of the Brain: Past Imperfect, Present Indicative—and Future Perfect?" [33] as well as in "Neurochemistry and Serotonin: A

Chemical Fugue" [34]. I need go no further here, because by that time neurochemistry was

well launched. Knowledge of serotonin soon accelerated at such a rapid rate that many were unable to keep abreast, and I was one of them. A new generation of experts has grown up, highly skilled technically and fully able to continue the rapid advance in progress. I would, however, remind them that there are large numbers of serotonergic neurons in

the central nervous system and that their functions still have not been integrated into the cholinergic and adrenergic neurons. The simplistic

way my generation looked at such substances as angiotensin, prostaglandins, and serotonin and its derivatives is gone forever. Gone too are

the days when a biogenic substance had only one function in the body. Serotonin has so many that have already become established that it makes one wonder where it will end. And this does not take into consid-

eration the large number of potential derivatives, as seen, for example, Perspectives in Biology and Medicine · Autumn 1976 | 7

in the melatonin of the pineal gland. The participation of serotonin in the mechanisms of sleep, of mood, and of arterial-pressure regulation makes me realize that serotonin or any other physiologically active substance must somehow fit into an overall, highly integrated pattern which

makes the life of complicated developed creatures not only possible but responsive to an equally highly integrated environment in which the organism must live. REFERENCES

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