A powerful small molecule; believing in GABA


rosemary-1

GABA (γ-aminobutyrate) is the major inhibitory neurotransmitter in the mammalian brain. It contributes to the all-important excitatory-inhibitory balance of neuronal communication. Dysfunction of the GABAergic system is implicated in disease, such as epilepsy, insomnia, schizophrenia and anxiety disorders. The history of GABA, from when it was first discovered to when its power as an inhibitory neurotransmitter was accepted, is an interesting one. It is wrapped up in personal accounts of the scientists involved, and looking back at the winding path of acceptance is fascinating.[1],[2],[3]

The story starts with the discovery of a ninhydrin-reactive material in mouse brain tumours by Eugene Roberts and Sam Frankel in 1949.[4] GABA had been known since 1910, of course, and had been found in bacteria and fungi and potatoes.[5] Once the structure of this ninhydrin-reactive material was confirmed to be GABA by Sidney Udenfriend, Roberts scribbles “the brain is like a potato!” in his notebook, and sets about proving its biological significance. [6],[7]

Roberts and Frankel’s work was published in 1950 for a conference at which Jorge Awapara also reported the presence of an “unidentified amino acid” found only in the brain.[8] By the time of the conference Awapara had also recognised it was GABA. At the conference a room-shortage led these two into close proximity, and Awapara and Roberts apparently agreed that Roberts would continue working on metabolism of GABA, and Awapara would continue working on the metabolism of taurine.7b

Six years pass, and scientists were still questioning whether the presence of GABA in the brain meant it had a role in signal conductance.[9] It should be noted that the debate over whether synaptic transmission was electrical or chemical had only just reached its chemical conclusion.[10] Ernst Florey had found Factor I, a chemical of unknown composition, extracted from mammalian brain and which inhibited the crayfish muscle system amongst others.[11] With help from Merck scientist Alva Bazemore, they purified over 45 kg of cow brain, and, using over 450 L of acetone they managed to yield crystalline Factor I. It was GABA, obviously.[12] There were still things to clear up, however, as some of their earlier work on the spinal cord of cats was initially unrepeatable by David Curtis, a fact now anecdotally put down to the allosteric modulation by the barbiturates also used in the experiments, or contamination of glycine in Florey’s earlier samples.11, [13]

Evidence against GABA being a neurotransmitter was accruing. The fact that GABA was so widespread in the brain was a worry for many in the field. “It’s just a metabolic wastebasket” remarked one great neurochemist.7b Strychnine was known to inhibit transmission in the spinal cord, and yet did not compete with GABA, which was another point of confusion at the time. With hindsight we know it was the as-yet-undiscovered glycine receptors in the spine causing havoc here.

Some scientists remained convinced there was more to the story, and Eugene Roberts was one of them. “Stop bringing your GABA solutions to our labs,” pleaded biologists, daily. “They take up room, and we’re not going to test them.”

In 1959 a notable conference was convened to hear the latest research and clear up the confusion. This proceeding (the 1st Interdisciplinary Conference on GABA) at a nearby pub (City of Hope Research Institute) was attended by many of the leading people in the field. Excitement was in the air as scientists from Australia, Canada, England, France, Hungary, Japan, the United States and the former Soviet Union bustled together, spilling pints and talking Neuroanatomy. Most researchers had a top night, and left feeling like they had made life-long friends and collaborations from all over the world.

GABA itself, however, fared badly. It was not going to join the echelons of the neurotransmitters quite yet. Alongside everything else it was seen not to have a rapid, enzymatic degradation pathway, unlike the newly-discovered acetylcholine neurotransmitter. This was thought to preclude it from involvement in rapid on-off communication. GABA was a metabolite with some generalised depressant nature, but that was all.[14]

It was not until nearly ten years later that unequivocal proof was published by Krešimir Krnjević and Susan Schwartz in 1967 that GABA was shown to be the major inhibitory neurotransmitter.[15] Intracellular recordings, as well as pharmacological tests revealing selective block by picrotoxin and bicuculline, clearly demonstrated GABAergic transmission.[16] After this proof, there was an exponential growth in GABA research. Writing in 1974, Eugene Roberts says ‘In the 25 years since the discovery of GABA … the status of the compound has passed from that of a biochemical curiosity and physiological enigma to that of a major inhibitory transmitter.’[17] At fifty years GABA receptors are shown to have decisive roles in many diseases, and they become big targets in drug discovery, including big-hitting benzodiazepines and general anaesthetics.[18] At 67 years, there are a huge 74,000 references containing ‘gamma-aminobutyric acid’ on Scifinder Scholar.[19]

The mixture of personal and peer-reviewed literature accounts of how GABA came to be accepted is what makes it fascinating. It is useless to deny that the direction of any scientific progress is open to contemporary influences and personal preferences. Perhaps it just reminds us that, as Carl Sagan writes: “Nature is always more subtle, more intricate, more elegant than what we are able to imagine.” And that means the path to understanding this transmitter will never stop winding.

Blog written by Rosemary Huckvale

POST SCRIPTS

  1. Dr Eugene (Gene) Roberts died last year on 8th November 2016 at the age of 96. His obituaries can be found here.
  2. This blog hopes to relate only a small portion of the history surrounding the establishment of GABA as a major inhibitory transmitter. There will have been other important contributions not mentioned here.

REFERENCES

[1] Johnston, G.A.R, GABA Australis, some reflections on the history of GABA receptor research in Australia, Pharmacological Research, article in press, 2016.

[2] Avoli, M., Krnjević, K., The Long and Winding Road to Gamma-amino-butyric acid as neurotransmitter The Canadian Journal of Neurological Science, 43:219-226, 2016.

[3] Krnjević, K. How does a little acronym become a big transmitter? Biochemical Pharmacology, 68:1549-1555, 2004.

[4] a) Roberts, E. and Frankel, S. γ-aminobutyric acid in brain. Federation Proceedings 9:219, 1950. b) Roberts, E. and Frankel, S. γ-aminobutyric acid in brain: its formation from glutamic acid. Journal of Biological Chemistry 187:55-63, 1950.

[5] Steward FC, Thompson JF, Dent CE. γ-Aminobutyric acid. A constituent of the potato tuber? Science 110:439-440, 1949.

[6] Udenfriend, S. Identification of gamma-aminobutyric acid in brain by the isotope derivative method. Journal of Biological Chemistry 187:65-9, 1950.

[7] Roberts, E.. Gamma-aminobutyric acid, Scholarpedia, 2(10):3356, 2007. B) Roberts, E., The History of Neuroscience in Autobiography Volume 2, Edited by Squire, L.R, Academic Press, 350-395, 1999.

[8] a) Awapara, J. Detection and identification of metabolites in tissues by means of paper chromatography. Federation Procedings, 9:148, 1950. b) Awapara, J., Landua, A.J., Fuerst, R., and Seale, B. Free gamma-aminobutyric acid in brain. Journal of Biological Chemistry 187:35-9, 1950.

[9] Roberts E. Formation and utilization of γ-aminobutyric acid in brain. Edited by Korey S.R., and Nurnberger J.I., Progress in neurobiology. 1. Neurochemistry. New York: Hoeber-Harper, 1956;11-25.

[10] Eccles, J.C., The Synapse: From electrical to Chemical transmission, Annual Review of Neuroscience, 5:325-39, 1982.

[11] For overview see Florey, E., GABA: history and perspectives, Canadian Journal of Physiology and Pharamcology, 69(7):1049-56, 1991.

[12] Bazemore, A.W., Elliott, K.A.C., Florey, E., Isolation of Factor I, Journal of Neurochemistry, 1:334-9, 1957.

[13] Johnston, G.A.R, GABA Australis, some reflections on the history of GABA receptor research in Australia, Pharmacological Research, article in press, 2016.

[14] Bowery, N.G, Smart, T.G, GABA and glycine as neurotransmitters: a brief history. British Journal of Pharmacology, 147:109-119, 2006. And references therein.

[15] a) Krnjević, K. & Schwartz, S. Is γ-aminobutyric acid an inhibitory transmitter? Nature, 211:1372, 1966. b) Krnjević, K. & Schwartz, S. The action of γ-aminobutyric acid on cortical neurones. Experimental Brain Research, 3, 320–326, 1967.

[16] Curtis, D.R., Duggan, A.W., Felix, D., Johnston, G.A.R., GABA, Bicuculline and central inhibition., Nature, 226:1222-1225, 1970.

[17] Roberts, E., γ-aminobutyric acid and nervous system function – a perspective, Biochemical Pharmacology, 23:2637-2649, 1974.

[18] GABA in the Nervous System: The view at Fifty Years, Edited by Martin, D.L and Olsen, R.W., Lippincott Williams & Wilkins, 2000.

[19] Accessed Jan 2017.


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