Discovering the neuron: from Golgi’s “black reaction” to Sherrington’s synapse
The discovery of Golgi’s ‘black reaction’ and its implications for observing nervous tissue had enormous scientific significance. From the improvement of it, modern research began on the histological structure of the nervous system and the units of which it is composed.
Eliana Berra – OPEN SCHOOL Cognitive Studies Milan
Advertising message So he remembered in his writings a Spanish doctor recalling his first observation, which took place in the rudimentary laboratory set up at a colleague’s home, of the nervous tissue treated with the “black reaction” created a few years earlier by the Italian Camillo Golgi.
The Spanish doctor was Santaigo Ramon y Cajal. The one who would have shared, with Camillo Golgi, the Nobel prize for medicine of 1906. The one who would have identified the essential building block of the nervous tissue in the neuron, bringing to the fore the cellular theory of the nervous system.
The “cell theory”, according to which cells are the elementary components of living organisms, was proposed in 1600, but was formalized only in 1800 by botanist Matthias Schleiden and zoologist Theodor Schwann and further validated, as regards the organs that make up the human body, by the German pathologist Rudolph Virchov. Until the beginning of the 1900s, however, it was debated whether this theory was also applicable to the nervous system; in fact, the nervous tissue appeared structurally more complex than that of other organs and the investigation methods of the time did not allow to distinguish the cells from the nerve fibers. What prevailed was the theory that the nervous system was the set of a dense network of thin filaments that joined together to form the nerve fibers, connected to each other. This theory, described by the German Josef von Gerlach in 1871, took the name of “reticular theory” and was embraced by most of the scholars of the time. Among them was Camillo Golgi.
Camillo Golgi graduated in medicine in 1865 at the University of Pavia under the guidance of a professor whose fame is still known: Cesare Lombroso. However, histological studies on nervous tissue began for the new graduate Golgi, when he joined the Pavia laboratory directed by Giulio Bizzozero, and would have stopped shortly thereafter if his determination had not prevailed and circumvented the difficulties he faced. In fact, he was appointed primary at a provincial hospital, the Pie Case degli Incurabili of Abbiategrasso, where research was not foreseen, there was no laboratory and the means available were rudimentary.
Golgi wanted to study the nervous tissue and to do so he wanted to observe the brain tissue as until then it had not been possible. Optical microscopes, used for scientific research since the 1600s, had developed further in the 1800s, but were still spoiled by some optical and chromatic artifacts. Furthermore, in order to observe the nerve tissues under the microscope, they had to be sectioned into very thin “slices” and treated with fixatives, which at the time were mainly alcohol and chromic acid, and dyes, such as carmine. However, these techniques did not allow for optimal results. To revolutionize the observation of the nervous tissue would have been precisely the “black reaction” of Golgi, “recipe” developed following the numerous attempts made in the kitchen / laboratory of Abbiategrasso.
recalls the researcher from Pavia in his writings. It was not enough. Only after numerous tests, Golgi understood that he could obtain the desired result by immersing the nervous tissue in a solution of chromium and, successively, of silver nitrate. Under the microscope, it was finally possible to observe the cells and nerve fibers, which stood out with their black profile on a light background.
A number of legends have spread about this discovery, according to which the “black reaction” was the result of an extraordinary stroke of luck. Some sources say that the researcher, with his elbow, accidentally spilled the silver solution on the brain tissue samples; others, that an attendant accidentally threw a sample of brain tissue into the trash where a sample of silver nitrate had been thrown a few hours earlier. In both cases, Golgi would have decided to reuse the samples equally, observing with great amazement the spectacle that was revealed to his observation under the microscope. It is not possible to be sure of the truthfulness of these episodes which, however suggestive, are quite unlikely. What is certain is that, regardless of how this discovery actually happened, its scientific scope was enormous: no other method of the time allowed such an observation of the nervous tissue. From the improvement of it, modern research began on the histological structure of the nervous system and the units of which it is composed.
Announced for the first time in 1873, the discovery of the “black reaction” was described in more detail the following year in the Italian Medical Journal.
Although Golgi was the first to have the opportunity to distinctly observe the black-stained nerve cells and their ramifications, which only decades later would have been baptized with the name of axons and dendrites, he drew some erroneous conclusions. With the limitations of the method, the dendrites and axons were clearly visible, but they seemed to form uninterrupted intertwining, without any solution of contiguity with each other. It was, for the Pavia researcher, a confirmation of the reticular theory of the nervous system.
To give a change, affirming the individuality of the nerve cells as constitutive elements of the brain tissue will be the Spanish researcher Cajal, who for many years will become for Golgi an opponent, albeit esteemed, in the European scientific panorama.
Cajal, a doctor returning from a military experience in the Cuban war, returned to Spain in 1875 and just then began to devote himself to scientific research. Characterized from a very young age by a creative, impulsive and passionate disposition, he decided to buy the necessary equipment for laboratory work with military pay, dividing in the following years between the universities of Madrid and Barcelona.
recalls Cajal in his writings. It was in 1887 that, for the first time, in the rudimentary laboratory set up in the home of a colleague and friend, the psychiatrist Dr. Simarro, he observed some samples of nervous tissue treated with Golgi’s black reaction.
Starting from that observation, Cajal began to use Golgi’s method in his laboratory, gradually making changes. The duration of immersion of the tissue in the solution varied according to the nervous structure it wished to study and the characteristics of the animal to which the tissue belonged.
It was thanks to these changes that he could observe the nervous tissue with an even greater definition. Thanks to his artistic talent, hindered in his youth by the family who nourished for him the aspiration of a medical career, he was able to faithfully reproduce what he observed under the microscope in hundreds of splendid drawings made by hand. With the improvement of Golgi’s “black reaction”, Cajal observed that some axons, although very close to the contiguous ones, ended freely, without direct connection with other nerve fibers. In 1889, the researcher concluded that nerve cells, as well as those of other tissues, were independent units. It was the confirmation of the cellular theory of the nervous system.
Advertising Message The results of his studies, however, were struggling to expand beyond Spanish borders. Therefore, in 1889, Cajal decided to participate in a prestigious congress in Berlin, paying the costs out of his own pocket since the university refused to finance it. Among the organizers of the event, there was an authoritative scholar of the time: Wilhelm von Waldeyer, director of the anatomy institute of the University of Berlin. Impressed by Cajal’s studies, Waldeyer dedicated himself to conducting a review of the research carried out up to then on nerve cells. The result was the publication in 1891 of a long work in six parts, in which, for the first time, the most important cells of the nervous system were baptized with the name with which we know them: neurons. They were defined as elementary and independent units from each other. Cell theory, thanks to Cajal’s studies and Waldeyer’s work, became the “neuron theory”. Soon, the branches of nerve fibers that originated from the body of the neuron also had a name: Wilhelm His named in 1890 with the name of “dendrites” the fibers that conduct the nerve impulse from the periphery to the cell body; in 1896 Albrecth von Kolliker called “axons” those that lead him from the cellular soma to the periphery. Despite the evidence supporting the neuron theory and the broad acceptance by the scientific community, no fierce opponents continued to oppose it. Golgi, who in the meantime had provided other authoritative and multifaceted contributions to scientific research, describing glial cells such as astrocytes,
The diatribe did not subside even when it was announced that the two antagonistic scholars, Golgi and Cajal, would share the Nobel Prize for medicine, one for the invention of the “black reaction” method, the other for exploiting it by establishing the structure and function of the neuron.
Cajal will comment in his writings. The ceremony took place in 1906, the same year in which the Italian Giosuè Carducci also received the Nobel prize for literature. Even this occasion became a pretext for the two scholars to defend their theories, launching attacks on the antagonist in their respective thanksgiving speeches.
If the cell theory of the neuron was now predominant, there was a big question that had yet to be answered. If nerve information travels along dendrites and axons and these are not joined together, how can the signal pass from one neuron to another in a short time, transferring information even over long distances? The reticular theory, by proclaiming the continuity of the nerve fibers with each other, had an advantage over this aspect. Both, however, did not answer another thorny question: how does the nervous signal originate and propagate?
To answer both questions, it would have been another pair of scholars who, ironically, also shared the Nobel Prize for medicine in 1932: they were the British Edgar Douglas Adrian and Charles Scott Sherrington. The first identified in electrical activity the mechanism underlying the transmission of the neuron’s nerve impulse, the second clarified how this impulse was transmitted between two or more neurons.
The idea that electrical activity represents the mode of transmission of nerve signals was already present since 1700 and followed the long Italian scientific tradition of scholars such as GianBattista Beccaria, Luigi Galvani, Leopoldo Nobili. Their theories, however, were largely criticized and ignored by other authoritative European researchers for over two centuries. It was necessary to wait until 1928 so that, thanks to Adrian, electrical activity could be legitimately recognized at the basis of the nervous impulse. In his famous experiment, the British doctor isolated a few axons from a nerve in a rabbit’s neck and placed an electrode in contact with them. The electrode, whenever the rabbit gave a breath, recorded electrical activity, which was converted into a rattle-like sound signal by an amplifier. Each rattle corresponded to the electrical impulse used by the neuron to transmit the signal and “dialogue” with neighboring neurons: the action potential. Today we know that the action potential is created because each neuron has an ever-present electrical charge, the “resting potential”. The resting potential is guaranteed by the structure of the neuron which, like a battery, presents on one side a positive electric charge outside the membrane that covers it, and on the other a negative charge, inside the cell. The positive or negative charge is given by the prevalence of electrically charged atoms, the ions, between the two sides of the membrane. When a stimulus comes to the neuron, there is a change in the concentration of ions from the two sides of the membrane and, consequently, in the electric charge. In particular, if the stimulus is excitatory, the difference in electric charge is reduced: thus there is the phenomenon called “depolarization”. If a certain “threshold” value of depolarization is exceeded, there is the “action potential”: electrical manifestation of the nerve impulse, it represents the way in which neurons spread their messages.
In a subsequent study on the toad, Adrian recorded the electrical impulses, amplified them and converted them graphically, visualizing them as pointed “peaks”. Observing the peaks, he discovered the action potentials of a given neuron were all equal in amplitude and duration, regardless of the intensity of the stimulus: what varied was their frequency.
Adrian therefore described how the nerve impulses, thus generated at high or low frequency, propagate along the axon, starting from the cell body of the neuron to its end, “like a flame along a lit fuse”. The “fuse” travels the axon along its entire length and can also travel for rather long routes. However, if as supported by the neuron theory, axons are not joined to each other, how is their propagation between different neurons possible?
The answer will be provided by what for many is recognized as the “nervous system philosopher”, Charles Scott Sherrington, and has a specific name: synapses.
Synapse, a term used for the first time by the English doctor, means “junction”, “union”, and represents the functional connection between two neurons through which the nervous signal is transmitted. At the time of Sherrington, there was talk of a “functional” structure since the existence of the synaptic space was confirmed and structurally observable only in the following decades, following the advent of the electron microscope.
At the synapse level, neurons communicate with each other through an amazing transformation of the message, which from an electrical impulse becomes a chemical signal.
As had happened with cellular and reticular theory, the electrical and chemical theory of the nerve signal were long debated and seemed irreconcilable to most scholars.
To reconcile these theories would have been the scientific contribution of a third pair of scientists and Nobel laureates in medicine in 1936: Otto Loewi and Henry Dale. Loewi’s experiments on the heart of a frog showed how an electrical stimulus, carried along the axon, was followed by the release of a chemical substance capable of transmitting the “message” between two neurons and between neuron and muscle with tangible consequences: the heart as a frog, depending on the electrical stimulus and the chemical released accordingly, it accelerated or slowed its beat. Henry Dale identified two substances involved in synaptic communication: norepinephrine and acetylcholine. They were termed neurotransmitters because of their role in transmitting information in nerve tissues. Afterwards, between the 30s and the 50s, other neurotransmitters were identified and their functions clarified: glutamate and glycine, with an excitatory effect; serotonin, involved in the mood; gamma-amino butyric acid (GABA), inhibitory; dopamine, involved in the circuit of pleasure and movement. The signals, which travel through different brain circuits involving numerous neurons and different synapses, are conveyed through a sequence of electrical impulses and neurotransmitters, which, as in an algebraic sum in cascade, add up or subtract from each other, in a myriad of different configurations. involved in the circuit of pleasure and movement. The signals, which travel through different brain circuits involving numerous neurons and different synapses, are conveyed through a sequence of electrical impulses and neurotransmitters, which, as in an algebraic sum in cascade, add up or subtract from each other, in a myriad of different configurations. involved in the circuit of pleasure and movement. The signals, which travel through different brain circuits involving numerous neurons and different synapses, are conveyed through a sequence of electrical impulses and neurotransmitters, which, as in an algebraic sum in cascade, add up or subtract from each other, in a myriad of different configurations.
In the decades following these discoveries, new studies have further clarified the mechanisms that regulate the communication between neurons and their circuits at the level of the nervous system, highlighting, on the other hand, an unexpected complexity, which has not yet been fully deciphered .
Sensing the charm and grandeur of this complexity, Sherrington himself wrote poetically: