70 Years on: the Platinum Anniversary of the Solution to Milk Ejection by the Mammary Gland. Part 3: What do Myoepithelial Cells do in other Exocrine Glands?

The German histologists of the 19th Century described myoepithelial cells in a number of exocrine glands including salivary, sweat and lacrymal glands. Later classical histologists and electron microscopists workers added to the list; snake venom glands are one example.

Jim Linzell, using the silver staining techniques that revealed the mammary myoepithelium, found myoepithelial cells in the sweat glands and submaxillary salivary gland. He remarked that up to the 1950s that the greatest interest in myoepithelial cells had been shown by pathologists, with respect both to the rare carcinomas they give rise to or their in metastasis. Looking at the more recent literature that generalisation still holds. The question is, though, what do the myoepithelial cells do in glands, other than in the mammary gland where they push milk stored in the alveoli into the duct system?

Top (8) Myoepithelial cells lying outside
cells of sweat gland in dermis. Cat.
Bottom (10) Submaxillary salivary gland
of cat showing smaller, less numerous
myoepithelial cells.
From Linzell, 1952.

The myoepithelial cells seem smaller in these other glands than in the mammary gland so the question arises: do they have any role in emitting secretion?

One function in the mammary gland I have not mentioned in the previous two articles is in relation to the mammary ducts. As well as the stellate or ‘basket’ cells which are arranged around the secretory alveoli, there are myoepithelial cells arranged longitudinally along the ducts. These, Linzell demonstrated, when stimulated to contract by oxytocin serve to shorten and widen the ducts. Obviously, milk will flow more rapidly in a shorter, wider vessel than in a longer, thinner one and so the two types of myoepithelial cell act in concert to move milk toward the sucking young.

There is evidence that the flow of saliva can be augmented under certain circumstances and that the contractile myoepithelial cells are be responsible. The nervous control of salivary secretion is complex and there remain many unanswered questions and anomalies in experiments dating from the 19th and first half of the 20th Centuries. Although written nearly 70 years ago, the best discussion I have found on this topic is that in the Physiological Society monograph, Physiology of the Salivary Glands, written by A.S.V (now Sir Arnold) Burgen and Nils Emmelin (1914-1997) published in 1961. Later in the 1960s, Nils Emmelin provided very strong evidence that the contractile myoepithelial cells, under nervous control (in contrast to the mammary gland), are involved in augmenting the rate of salivary secretion.

The strength of mammary myoepithelial contraction can have surprising consequences. Many infants—and passing adults—have been shot in the face by a stream of milk from their mother. Such streams formed the basis of Tintoretto’s famous painting The Origin of the Milky Way which can be seen in the National Gallery in London. Greek myths are all double Dutch to me but the illustration is of Hera who was suckling Heracles; he sucked* so strongly that Hera pushed him away. Her milk shot out in all directions (remember each opening in the human nipple is from a separate gland) across the heavens and formed the Milky Way^†.

The Origin of the Milky Way
Tintoretto

S.J. Folley (see part 1 of this series) used that painting to illustrate the depiction of milk ejection in mythology and art in his Dale Medal Lecture to the Society of Endocrinology in 1969. Folley had not played any great part in discovering or delineating the milk-ejection reflex (although he had encouraged those who did) but it remained an abiding interest. I was in the audience at that lecture and it would not have been possible to guess that Folley was virtually completely blind and that he had committed to memory the whole script complete with hand gestures that pointed to the slides^‡.

In some snake venom glands, ejection of stored secretion is achieved by external muscles pressing rapidly on the gland to eject the secretion at high pressure. Whether or not myoepithelial cells are involved in these front-fanged snakes or in the low-pressure ejection system of rear-fanged snakes appears not to have been studied. Obvious questions are: are the myoepithelial cells innervated? Do they respond to mechanical stimulation such that they contract as prey is being held tightly by the jaws and thereby help squeeze venom out of the gland of rear-fanged snakes?

I hope I have illustrated that 70 years after they hit the headlines there are many basic questions that remain on the rôle of myoepithelial cells and, indeed, on how exocrine glands work, and that in the Gadarene swine-like rush into molecular biology whole areas of biological organisation have been abandoned. 

*The entry in Wikipedia gets suckling wrong, as if often the case. Mothers suckle; babies suck is the accepted distinction. Mammals suckle their young. Suckle: to give suck to. Roast sucking pig not suckling pig etc etc.

†There was a famous exchange between Alec Bangham (1921-2010) and Jim Linzell (1921-1975) at Babraham. Alec Bangham then working on his later famous liposomes said to Jim who was working on lactation in goats, James, you will never get into the Royal Society on a ruminant’s back. Ah, said Jim, but there is always the Milky Way. It is an open secret that Jim was on the list for election in 1976 when he died in December 1975. Alec was elected in 1977.

#At a Gordon Conference in 1975, Jim Linzell was giving the main invited (Thursday Night) talk. Folley had recently. When Linzell came to a problem to which he didn’t have an answer, he looked towards the heavens and said, Perhaps Folley knows. Instantly, Howard Bern proclaimed loudly, He’s looking in the wrong direction!

Burgen ASV, Emmelin NG. 1961. Physiology of the Salivary Glands. London: Edward Arnold.

Folley SJ. 1969. The milk-ejection reflex: a neuroendocrine theme in biology, myth and art. Journal of Endocrinology 44, 476–90. 

Weinstein SA, Smith TL, Kardong KV. 2009. Reptile Venom Glands. Form, Function, and Future. In, Handbook of Venoms and Toxins of Reptiles, edited by Stephen P. Mackessy, pages 65-91. Boca Raton: CRC Press.

70 Years on: the Platinum Anniversary of the Solution to Milk Ejection by the Mammary Gland. Part 2: James Lincoln Linzell

Jim Linzell, June 1974
Photographed at Babraham by
Alec Bangham

It was in May 1948 that Jim Linzell (1921-1975) realised he had, just like Keith Richardson, stained myoepithelial cells while looking for nerve fibres in the mammary gland.

Linzell had escaped veterinary practice and was working for a PhD in the University of Edinburgh’s physiology department. He had qualified as a vet in London but found veterinary practice frustrating and unrewarding in that he lacked the time to be thorough while having to undertake jobs that technicians were perfectly capable of doing. He changed direction by going to Edinburgh to work under Professor Ivan de Burgh Daly FRS (1893-1974). He was funded by an Agricultural Research Council (now morphed into the BBSRC) research traineeship. He told Daly that he would like to work on the uterus and placenta but Daly told him to find out everything about the anatomy and physiology of the mammary glands in as many species as he could; this he did until he died at the age of 54.

Linzell did not like Edinburgh. Cold, dank and unfriendly was how he described it. Jim and Audrey, together with their two sons, lived in a flat relatively near the university. He published a note in Veterinary Record which gave his address: 7 Blackwood Crescent. His trenchant views on that flat, the landlord, the neighbours and the inhabitants of Edinburgh were expressed many times in later years whenever Edinburgh was mentioned. He would be horrified by its current valuation of £210,000.

The Linzell family had
a flat in this house in
Edinburgh
Google Street View

But Jim was not stuck in Edinburgh. A great deal was happening to Daly and the future of agricultural research. During the Second World War it was realised that the lack of progress in increasing food production was caused by lack of knowledge of how animals work. To this end, Daly was given the job of setting up a new research institute. He found a run-down stately home for sale near Cambridge and set about building laboratories, animal houses and housing for staff. Staff were appointed ahead of space becoming available and Linzell became one of the first members of staff of what is now the Babraham Institute on 1 October 1948, albeit based for a time in Edinburgh. Then, a temporary measure, Daly converted rooms in Babraham Hall into laboratories and a wash-house to hold Linzell’s goats. Linzell was one of the first scientists on site in 1950 as Daly eviscerated his old department in Edinburgh by appointing members of staff to Babraham posts.

It was in Edinburgh that Linzell found he had revealed myoepithelial cells in the mammary gland of cats. He was studying the role of nerves in controlling blood flow by classical physiological and histological techniques. In his first paper, published in Journal of Anatomy in 1952—Jim never rushed into print—he confirmed all the findings of Richardson. He also tried various methods of silver staining which were all notoriously capricious but extended his work beyond one species. He showed clearly the presence of myoepithelial cells in cat, dog, rabbit, rat, goat, and human mammary glands.

An important additional finding was that the myoepithelial cells have no connexions to the nervous system, thereby refuting one suggestion that the milk-ejection reflex was a pathway composed entirely of nerves.

He then went further. By observing the living gland he saw contraction of the myoepithelial cells in mouse, rat, guinea pig and rabbit and was able to study what caused them to contract and thereby expel milk. Oxytocin dropped onto the gland, of course, worked. So did direct electrical stimulation was one would expect with a muscle but stimulation of nerves in the region had no such effect. Sometimes, light mechanical stimulation was enough to do the trick.

Linzell’s observations provided the final link in the chain for the neuroendocrine milk-ejection reflex.

Two figures from Linzell's 1955 paper. On the left the alveoli can be seen
full of milk. When oxytocin was dropped on the gland (right) the alveoli
contracted and milk was driven into the duct (D)

Linzell’s observations on the living myoepithelium were presented to the Physiological Society at its meeting of 18-19 December 1953. That was his first appearance before the Society, a year after his election as a member. Presenting a paper was a daunting experience because right through the 70s and into he 80s there was a phalanx of Nobel prizewinners in the audience. Discussion was often fierce and every word in the circulated abstract had to be agreed before a vote was taken on whether or not the paper should be accepted for publication in Journal of Physiology. Because the paper was refereed by attendees it could be included in reviews that demanded reference only to refereed papers. For some reason I do not understand or agree with that system was dropped by the Physiological Society in the early years of the present century. Sometimes, if all the material had been published in the Proceedings, it was not necessary to clutter the literature with a full paper. However, Linzell had covered a lot of ground in his observations and a full paper appeared in Journal of Physiology in 1955. 

Neither Keith Richardson nor Jim Linzell took any further part in work on the milk-ejection reflex.

In the past I have used the history of the milk-ejection reflex to illustrate the fact that many advances in human physiology and medicine have come from fundamental research funded as part of agricultural research.

But there remain many unanswered questions about myoepithelial cells in exocrine glands—not in the mammary gland but elsewhere in the body.

Linzell JL. 1952. The silver staining of myoepithelial cells, particularly in the mammary gland, and their relation to the ejection of milk. Journal of Anatomy 86, 49-57.

Linzell JL. 1954. The contractility of the alveoli of the mammary gland. Journal of Physiology 123, 32P.

Linzell JL. 1955. Some observations on the contractile tissue of the mammary glands. Journal of Physiology 130, 257-267.

70 Years on: the Platinum Anniversary of the Solution to Milk Ejection by the Mammary Gland. Part 1: Keith Clifford Richardson

Seventy years ago, the final bricks were being put in place that established the milk ejection reflex, a classic neuro-endocrine pathway. At the start of suckling receptors in the nipple or teat send nervous impulses to the brain. Nervous pathways in the brain then induce the release of the hormone oxytocin from the posterior lobe of the pituitary gland. Oxytocin is then carried in the blood to the mammary gland where it causes contraction of myoepithelial cells that surround each secretory alveolus containing stored milk. The milk is thus forced into the duct system of the gland where it can be removed by the sucking infant.

My old lecture slide showing the milk ejection reflex - the pathway shown
in white

The Problem

Although evidence for that reflex had been obtained there was still confusion as to which cells in the mammary gland oxytocin was acting on. The description of myoepithelial cells (and their probably contractile nature) by a string of famous German histologists began in 1850. By the end of that century, it was established that there were two types of myoepithelial cell in the mammary gland: stellate around the secretory alveoli and spindle-shaped along the outside of the ducts. However, those working on milk ejection in the first half of the 20th Century were apparently unaware of the earlier work by the German histologists and got into what can only be described as a right mess. They confused myoepithelial cells with smooth muscle, with capillary adventitial cells and the basal cells of the mammary epithelium. By the 1940s, the general view seemed to be that the very sparsely distributed smooth muscle in the mammary gland was responsible.

In the late 1940s, two people working independently, sorted the whole matter out, because of the same serendipitous finding. Their results were in total agreement and with follow-up physiological studies by one of them, the whole story was complete.

Keith Richardson in London

Keith Clifford Richardson (1905-1997) was the first to do the work and the first to publish in 1949, although by the time of publication Jim Linzell (James Lincoln Linzell, 1921-1974) had completed the histological phase of his research (published in 1951).

In 2009 the late Isabel Forsyth (1936-2016) and Peggy Neville included a short account of how Richardson came to be interested in the mammary gland in their introduction to a volume of Journal of Mammary Gland Biology and Neoplasia. He was asked for assistance as a histologist by Sydney John Folley (1906-1970) of England’s then National Institute for Research in Dairying who was working on the hormonal control of mammary development and lactation in goats. Isabel, who worked in Folley’s department, reported that Richardson was, at first, somewhat reluctant since he found himself responsible for moving the Department of Anatomy of University College London back to its pre-war premises in Gower Street from its wartime home in Surrey. Eventually, however, he agreed. A long correspondence with Folley (they were of similar age) ranged from science to their mutual interest in modern art.

Both Richardson and Linzell were trying to stain nerves in the mammary gland. Both realised that the silver impregnation technique revealed the myoepithelial cells. Richardson argued that to show their contractile nature it would be necessary to obtain sections from both full and empty glands. This he did by perfusing goat mammary glands through the artery with fixative.

Richardson’s summary expresses perfectly what he found:

Myoepithelium covers the stromal surface of the epithelium of the alveoli, ducts and cisterns of the entire gland, and is thus much more abundant than is generally realized. Smooth muscle forms scattered inter-lobular bundles closely associated with the blood vessels. The theory that myoepithelial contraction is the principal factor con­cerned with ‘let-down’ and the ejection of milk is examined; other factors such as inter-lobular smooth muscle contraction, vascular changes, and elastic recoil of the stroma appear to play minor roles, if any, in this phenomenon. Hitherto, it has been assumed that myoepithelial cells are contractile because they bear structural resemblances to smooth muscle fibres. With the new technique structural changes have been found in the myoepithelium of contracted as compared with distended alveoli and ducts. These changes, together with the general orientation of myoepithelial cells, and the precise relationship between these cells and the folds in the secretory epithelium from contracted glands, are consistent with the assumption that myoepithelium is the contractile tissue in the mamma which responds to a neurohormonal mechanism involving oxytocin. 

Figure 12 from Richardson's paper: a small
contracted alveolus in surface view,
showing a myoepithelial cell with
nucleus n and branching processes

Figures 16 and 18. Same magnification
16: Myoepithelium on the surface of distended alveoli
18: Myoepithelium on contracted alveoli

Information on Keith Richardson is sparse. He was born on Christmas Day 1905 in Geelong, Victoria, Australia. He graduated from University of Western Australia in 1926, with an MSc following in 1927. As was often, indeed usually, the case at the time he did not have a PhD but whether he derided it as did many of his colleagues as ‘the German degree’ is not recorded. He was a lecturer in histology in UCL’s Department of Anatomy by the late 1930s. In the 1939 Register he was staying at the Grand Hotel, Westgate Street, Cardiff and was described as a histologist and university lecturer in the Department of Anatomy of UCL.

There are some recollections of Richardson by former students and colleagues.

Ray Guillery FRS (1929-2017):

Keith Richardson taught the histology classes during the first terms of the medical course. He was tightly organized throughout. Not only did he start and finish each lecture precisely on time, but every sentence was a complete and elegant structure. During this introductory course and for many years at UCL thereafter I learned from him a respect for order and precision. It was through the histology course that I was particularly stimulated to take an interest in anatomy. Looking back now this is perhaps surprising. The lectures were not as exciting as many others, nor as original, but they opened doors for a completely ignorant student who had not yet been taught the importance of recognizing originality, or creativity. I think Keith understood his audience. 

Ruth Bellairs:

Within the Anatomy department the third floor (the top floor at that time) was given over mainly to histology. At the south end the Reader in Histology, Mr Keith Richardson occupied the suite of rooms facing Foster Court. He was a cherubic-looking bachelor in his fifties who published little but whose high technical standards were legendary. He was a strict disciplinarian in his teaching and no chattering or whispering by the students was ever permitted in his practical classes. The north end of the third floor, now part of Physiology, was taken up entirely by the Histology teaching laboratory 

In an interview with Lynn Bindman (LB) for an oral history project of the Physiological Society conducted by Laurence Smaje (LS), Alex Cooper (AC) and David Miller (DM) at Lynn’s home in London on 30th March 2015, the conversation turned to Richardson:

LS: [Keith] Richardson. LB: Richardson, thank you. So he was in the Anatomy department and he gave nine o’clock in the morning histology lectures. LS: Starting exactly at nine and finishing exactly at five minutes to ten. LB: And I suppose they were dry but they were just fascinating because actually he gave the physiological function of every structure he talked about. So for the skin you knew why there were hairs and sweat glands and everything else. DM: These were large room lectures together with the medics, or together with other science...? LB: Yes. Science, medics and dental students. Well, the Richardson ones I think were in a smaller theatre, there were about 100 odd. LS: Oh dear. LB: I don’t know how... LS: Well, they were given in the anatomy theatre, I think. LB: Were they? I remember... LS: Well at least they were when I was there. LB: I remember Embryology theatre, actually, that’s my memory.LS: But he left, didn’t he, to go to the States. LB: Right. LS: Because they wouldn’t give him an electron microscope. LB: Right, how fascinating. But he was such a brilliant teacher.LS: He was, excellent. 

After mammary glands: Keith Richardson in the U.S.A.

And to the U.S.A. he did go. Shipping records show he visited the U.S.A. in 1956-57 but his move to the U.S.A. was in 1959. He left Southampton for New York on 25 February 1959 aboard the French liner S.S. Liberté (the former German ship Europa, received by France for war reparations). His U.S visa was issued in 1959. In 1960 and 1961 he can be found in St Louis as Associate Professor in Washington University Medical School’s Department of Anatomy. In passing it is interesting to note that in his 1949 paper he had, in modern parlance, rubbished the efforts of the chairman of that department, Ed Dempsey (1911-1975) in trying to distinguish myoepithelial cells from other cells in the mammary gland.

By 1962 Richardson was at the National Institute of Neurological Diseases and Blindness of NIH. 

His papers from his time in the American system involved electron microscopic studies of the nervous system, particularly the nature and storage and recapture of noradenaline (norepinephrine to U.S. readers) in the sympathetic nervous system. He collaborated and published with the Julius Axelrod of NIH who was awarded the Nobel Prize with Katz and von Euler in 1970. As remarked above, Richardson’s publications were few but of very high technical quality.

Floyd E. Bloom recalls Richardson at NIH:

I decided to see if I could learn some of the rudimentary methods of electron microscopy by becoming a guest worker in the NIH Campus laboratory of Professor Keith Richardson, head of the Section on Neurocytology. He agreed I could come over, and he walked me through the details of brain perfusion fixation, dissection for electron microscopy, orientation of the tissue blocks for embedding in epoxy plastics, and how to make glass knives from whole sheets of glass in order to do ultramicrotomy to acquire the less than 1000 Angstrom thick sections that could be placed on copper grids and examined in the electron microscope. Richardson had been the head research assistant in the neurocytology laboratories at University College London before coming to the NIH [He had been lecturer in histology]. Because he was one of the few Section heads who didn’t have a doctoral degree, he called himself “Professor Richardson” in the NIH Directory. (Julius Axelrod was another until he got his degree a few years before his Nobel Prize)…In addition to teaching me the fundamentals of electron microscopy, Richardson had been one of the first people to examine tissues of the peripheral autonomic nervous system. He had observed that immersion fixation with a mixture of osmium tetroxide and potassium dichromate produced sympathetic nerve fibers containing synaptic vesicles with dense granular cores, rather than the electron lucent synaptic vesicles that epitomized the nerve terminals at the neuro-muscular junction where acetylcholine was the recognized neurotransmitter. Then just before I left NIH for Yale, Richardson published a report in Science with Julius Axelrod and Lincoln Potter showing a second method for detecting norepinephrine by ultrastructural autoradiography of nerve terminals in the pineal exposed to 3H-norepinephrine; there the sympathetic nerves actively transported the norepinephrine into the nerve terminals, where it was stored in the synaptic vesicles. I was eager to test these approaches in the brain. 

Richardson’s later publications, possibly after retirement from NIH, were from the University of Maryland’s School of Medicine in Baltimore. I have found listed what appears to be text book for students, Illustrations of Light Microscopical Preparations from Various Tissues and Organs, published by the Department of Anatomy in Baltimore in 1976. The last publication I can found in PubMed was in 1977 when he would have been 72.

Keith Richardson must at some time after he retired return to London. He died in Camden on 15 February 1997, aged 91.

I have been unable to find a photograph of Richardson. If any reader knows of one or has further information please contact me.

Coming Next

Keith Richardson submitted his paper on 29 June 1948. A month previously Jim Linzell had  made his serendipitous finding that confirmed Richardson’s observations. In Part 2, I will look at Jim Linzell’s anatomical studies and how he then put the final brick in place by then observing living mammary myoepithelial cells in action.

Bloom FE. 2012. Floyd E Bloom. In, The History of Neuroscience in Autobiography, Volume 7, pages 1-56, edited by Larry R Squire. Oxford University Press.

Forsyth IA, Neville MC. 2009. Introduction: The Myoepithelial Cell and Milk Letdown; Entrance to the Multifunctional Role of Oxytocin. Journal of Mammary Gland Biology and Neoplasia 14, 221-222.

Guillery R. 1998. Ray Guillery. In, The History of Neuroscience in Autobiography, Volume 2, pages 132-167, edited by Larry R Squire. San Diego: Academic Press.

Richardson KC. 1949. Contractile tissue in the mammary gland, with special reference to myoepithelium in the goat. Proceedings of the Royal Society B 136, 30-45.

Carry on Cleo. Two unusual glands: the mammary gland and the snake venom gland

I usually try to avoid writing about mammary glands and lactation since I have been doing that for nearly fifty years. However, I cannot resist drawing attention to a question I have been asking for nearly that  long.

Most exocrine glands, salivary glands for example, do not store their own secretion. They simply switch on when a supply of saliva, say, is needed. By contrast, the mammary gland secretes milk continuously and stores its secretion internally until milk is removed at intervals. Another gland—or glands—that share that characteristic is the snake venom gland which builds up a supply of venom to be ejected on striking prey or in defence. When Cleopatra killed herself by grasping a viper to her bosom, two unusual exocrine glands were brought into close apposition.

Embed from Getty Images

We know that the rate at which milk is secreted is controlled by the volume of milk removed. The more milk taken, the faster the rate of secretion. In other words, there is a feedback mechanism operating to match demand by the young to supply by the mother. But what about the snake venom gland? There is every reason to expect that the rate of venom secretion would be controlled. Venom, like milk, is expensive for the body to produce and one might predict that a snake with a full load of venom would not continue to produce more only for it to be broken down again. So the questions are: 1. Does the rate of venom secretion increase after a strike? 2. Does the rate decrease as the internal storage compartment fills? 3. If so, by what mechanism is such control of secretory rate exerted? There is evidence that the answer to the first of these two questions is ‘yes’.

This is Scott A. Weinstein, Tamara L. Smith, and Kenneth V Kardong writing in their review, Reptile Venom Glands. Form, Function, and Future, published in 2009 (references deleted):

Venomous snakes hold stored venom during extended periods of fasting, but it remains ready when feeding resumes after hibernation or in defense; there is no reported turnover of the stored venom protein. If manually depleted (extracted, or "milked"), the secretory epithelium of the main venom gland exhibits rapid protein synthesis with subsequent exocytosis replenishing venom stores in the ductules and large lumen. This process is completed in about 16 days. However, when expending venom during natural strikes, venom is replenished more rapidly, or less total venom is expended as judged by the rapid recovery of lethal envenomation of prey.

The question (3)—not an easy one to answer by experiment—remains of how the mechanism to control the rate of venom secretion works. Is feedback control exerted chemically, like the mammary gland in full lactation, or physically by increased pressure stretching the secretory epithelium, as can be induced in the mammary gland by the sudden cessation of milk removal?

From Weinstein et al. 2009

Weinstein SA, Smith TL, Kardong KV. 2009. Reptile Venom Glands. Form, Function, and Future. In, Handbook of Venoms and Toxins of Reptiles, edited by Stephen P. Mackessy, pages 65-91. Boca Raton: CRC Press.