The trapezoid body, located in the brainstem, is part of the auditory pathway where nerve fibers from the cochlea on one side of the brain cross over on their way to the superior olivary nuclei to the other side, which functions in multiple aspects of hearing.  This crossing over is believed to help with localization of sound.  

Although modern-day diagrams generally show the nerve fibers originating on only one side of the brain for clarity, sections of the trapezoid body examined under a microscope display nerve fibers running in every direction.  Despite this, Cajal was able to clearly see the crossing of the nerve fibers from one side of the brain to the other and to correctly infer that signals ran from the ciliated cells in the Organ of Corti in the cochlea (B) up and across through the trapezoid body (E) to the superior olivary nucleus (J) on the opposite side of the brain, and from there up through the inferior colliculus (G) and towards the cerebral cortex.  

In addition, Cajal depicts some auditory fibers originating in the superior olivary nucleus as sending signals to the 7th cranial nerve (VII), otherwise known as the facial nerve, as well as to the 11th cranial nerve (XI), which controls muscles in the neck and shoulder.  He hypothesized that the connections of these nerves to the auditory system would thus explain the reflex displayed when one hears a sound and immediately turns one’s head and neck in the direction of the source.

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The olfactory bulb transmits odorant information from nose to brain through six layers of neurons: the olfactory nerve layer (not shown), the glomerular layer (A), the external plexiform layer (B), the mitral cell layer (C), the internal plexiform layer (D), and the granule cell layer (E), which is deepest in towards the brain.  Cajal definitively delineated the above six layers in 1890, building on earlier work by Schwalbe, and noted that scent information was not passed from single neuron to single neuron, as in other sensory systems, but was sent from the group of neurons comprising an olfactory fiber to multiple cells in the olfactory bulb.  

Cajal was also the first to observe that scent signals are not necessarily transmitted in a serial manner; the mitral cells project into the glomerular layer and are in contact with some ends of the olfactory fibers, and their output is sent to structures deeper within the olfactory cortex.  While he identified several types of cells present in the granule cell layer on a morphological and spatial basis, the function of these cells was not elucidated until recently: the provision of inhibitory feedback for mitral cells.

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Interstitial Cells of Cajal

Intestinal villi are small projections that extend into the lumen of the small intestine to increase the surface area of the intestinal wall, thereby increasing absorption of nutrients.  As the small intestine is made of smooth muscle, one might expect that it is innervated to enable consistent peristalsis.  

Cajal’s studies of the innervation of the small intestine revealed several types of cells that responded to the Golgi staining method he usually used.  Cajal believed that the stained cells were neurons, but his contemporaries such as Kolliker and Dogiel maintained that the stained cells in the intestinal villi were fibroblasts, as Golgi’s staining method was known to sometimes highlight connective tissue as well as nervous tissue.  

This controversy continued until the mid-1990s, when it was determined through the use of marker genes that these cells are generally of mesodermal origin, although some may originate from neural tissue.  Such cells are now identified through ultrastructural characteristics and are known definitively to serve as pacemakers for smooth muscle contraction.

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The cerebellum was the first system that Cajal studied, and it was through careful observation of its stained tissues that he first became convinced that neurons were distinct cells as opposed to part of a contiguous network, as well as one of the formative systems where he would develop his law of dynamic polarization.  The cerebellar granule cell is the most numerous type of neuron in the brain, constituting three-quarters of the neurons therein.  

Cerebellar granule cells possess four or five dendrites, each ending in a dendritic claw, a relatively large nucleus, and long, thin axons that split in two to form a “T” shape, forming a structure known as a parallel fiber.  Each granule cell gets input from mossy fibers, whose axons fit into the dendritic claws, and sends its output to Purkinje cells through its parallel fibers.  

This drawing by Cajal details the development of a cerebellar granule cell from its first appearance as a primary embryonic cell (1), through the beginning of its polar outgrowths (2, 3), formation of a horizontal bipolar cell (4), the start of its descending outgrowth (5, 6), its phase of vertical bipolarity (7, 8), its production of provisional dendrites (9, 10) and, finally, the pruning and refinement of its definitive processes (11, 12).  In more recent years, Cajal’s studies of the cerebellum have been corroborated through the use of techniques such as electron microscopy and histochemical staining, which have refined our understanding of the fine organization of the cerebellum.

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The fovea centralis, a small avascular depression at the center of the inner retinal surface filled with densely packed cone cells, is responsible for collecting visual information from the focus of visual gaze to form high-resolution images.  Signals from the fovea constitute half of all input to the visual cortex.  

In this drawing of the foveal pit (F) and perifoveal area by Cajal, the midget bipolar and ganglion cells (so called due to their small size) are highlighted.  In the image, each cone cell (A) contacts a single midget bipolar cell (B), which in turn makes synapses with a single midget ganglion cell (C), which then transmits information through the optic nerve to the visual cortex.  Cajal correctly noted that as distance from the foveal pit increases, the number of cone cells providing input to a single ganglion cell (C2) increases, a process known as convergence.  The lack of convergence at the foveal pit is what allows for maximum visual acuity.  

What Cajal did not realize was that, in order to maintain such visual acuity, each cone cell in the fovea is actually connected to two midget bipolar cells and midget ganglion cells (each pair of which exists in mutually exclusive planes of focus in microscope preparations) — one pair will signal only when the center of the cone’s photoreceptive area is stimulated, and the other pair signals only when the off-center area of the cone’s photoreceptive area is stimulated.  In addition, information transmitted from the fovea to the visual cortex also contributes to color vision.  

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The complicated circuitry of the brain and the precise connections of groups of neurons at various places within it suggest that neurons do not randomly form synapses as they grow, but instead respond to organizing signals.  Cajal was perhaps the first scientist to observe conical, fluid projections at the ends of developing neurons, and certainly the first to posit that such a structure might be involved in guiding the neuron towards a particular target.   

What we now know as growth cones – dynamic extensions of a developing neuron – respond to a variety of chemicals, secreted by target neurons and the extracellular matrix, that can be attractive or repulsive to the growth cone depending on the receptors present.  

The variety of the growth cones shown above is due to the complexity of the paths they were to navigate: the ones depicted in C were travelling a quick path through white matter in the brain, while those in A and B travelled a slower, more complicated path through gray matter and the ventral commissure, respectively.  

Current research has found that the shape of the growth cone in vivo differs somewhat from Cajal’s preserved specimens, but the complexity of the growth cone in relation to its travel speed remains the same.

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The Calyx of Held, first described by Hans Held in 1893, is one of the largest synapses found in the mammalian brain.  As part of the auditory system, each Calyx is part of the axon of a globular bushy cell in the anteroventral cochlear nucleus, which forms a synapse with a principal cell in the medial nucleus of the trapezoid body.  These synapses are integral to detection and localization of high frequency sounds.  

At a time when the existence of synapses between nerve cells were not yet accepted as fact, Cajal’s drawings of the mammalian auditory system revealed a sophisticated understanding of the relationship between the Calyx of Held and the neuronal cell body it envelops.  Rather than portraying[WC([1] them as a single, connected entity, Cajal’s coloring of this drawing indicates that he knew each part belonged to a separate cell, and that information would need to travel between them.  

Because of the large size of a Calyx of Held and its relative accessibility, it has been a popular model system for neurobiological research.  In particular, its amenability to patch-clamping has made it a favorite for systematic studies of presynaptic mechanisms, which can be involved in neurological disorders such as Parkinson’s disease.

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The neocortex, the complex strata of neurons in the mammalian cerebrum, comprises the ridges and grooves seen on the top layer of any 3-D depiction of the human brain. It is involved in processes such as sensory perception, language, and conscious thought.  

In the 1890s, Cajal and another scientist, Gustaf Retzius, independently identified bipolar neurons with horizontal nuclei in the developing neocortex of different types of mammals.  Shown in layer d of the drawing above, these cells – now known as Cajal-Retzius cells – are not only important for the transmission of information through the neocortex, but they also play a role in brain development.  They secrete a protein called Reelin, which is a critical component of a signaling pathway for neuronal migration, ensuring that new cortical cells migrating along the long radial glia (a) and into the upper layers of the neocortex will stop when they reach the correct position.  

Diseases associated with a lack of Reelin expression include schizophrenia, bipolar disorder, and autism.  Although Cajal was not aware of the existence of Reelin, or the developmental importance of the Cajal-Retzius cells, he accurately captured many of the major features of neocortical development using only his microscope.

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The olfactory system comprises three major parts: the olfactory epithelium, which contains olfactory sensory neurons in direct contact with the environment; the olfactory bulb, which integrates input from the olfactory sensory neurons; and the olfactory cortex, which processes input from the olfactory bulb is processed and transmitts information to other structures in the brain such as the thalamus and hypothalamus.  

By observing the positions of the axons and dendrites in relation to each other, Cajal was able to infer the direction of the flow of information in the olfactory system (represented above by arrows) from the periphery of the body to deeper brain structures, nicely illustrating his Law of Dynamic Polarization.  Cajal also noted that some information was transmitted back from the cortex to the bulb; he named this bidirectional information flow “centrifugal input”.  

The drawing above shows several olfactory sensory neurons (A) forming synapses with a single glomerulus cell in the olfactory bulb (B); in fact, olfactory sensory neurons expressing the same odorant receptor do converge on a pair of glomeruli cells, which integrate the multiple inputs before sending the signal onwards.  The olfactory sensory neurons are not necessarily spatially close, but Cajal seems to have found a lucky grouping in this drawing, illustrating again his talent for capturing structures with future significance.

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Insect eyes are compound eyes: each eye is made of many small unit eyes (ommatidia), as opposed to a single large unit eye as in vertebrates.  Cajal’s drawing above details the many ommatidia present in a single fly eye, as well as the neural circuitry relaying visual signals in towards the brain.  In the lamina complex, labeled C, we can see the axons of the photoreceptor cells crossing over each other, suggesting neural superposition.  

Neural superposition, discovered by Valentino Braitenberg and by Kuno Kirschfeld in 1967, is a phenomenon where photoreceptors from different ommatidia that receive the same signals converge upon the same synapses in the brain, providing vision that is both highly sensitive and high resolution.  Cajal was unaware of the phenomenon, but his drawings suggest that he recognized the organization of structures which make it possible.  

While visibly different from the vertebrate visual system, the insect visual system shares with it certain similarities in neural circuits: both systems have intermediary neurons that receive information from photoreceptor cells and direct it onwards towards the “neural switchboard” or neuropil.

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The vertebrate retina is another good example of Cajal’s Law of Dynamic Polarization.  Because he knew from which direction 

the stimulus to the photoreceptors (at the top of the diagram) originated, it was clear that information ought to be transmitted inwards, through axons of the photoreceptors to the dendrites of the horizontal and ganglion cells, and onward into the visual cortex, as noted by the arrows on the diagram above.  

In addition to the usual retinal information flow, Cajal illustrated some more unusual directions for information transmission.  He noted information flowing from c to I, along the cell body of a horizontal cell, although this subsequently has not been found to be a viable signal path.  Likewise, visual signals in many amacrine cells, shown to the right of D, need not pass through the cell body, contradicting the apparent intent of Cajal’s arrows. 

While Cajal may have erred in these two instances, he correctly observed another unusual signaling pathway between G and H: these axons from the visual cortex are returning to the retina and transmitting information to the amacrine cells.  This phenomenon has indeed been observed in present day, but not in all vertebrates; significant centrifugal input is a hallmark of avian eyes in particular.

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Once thought to be mere “filler” for the space around neurons, astrocytes are star-shaped glial cells found throughout the brain and spinal cord that are now known to perform many important functions, such as regulating the transmission of ions and glucose between blood vessels and the brain.  

When Cajal drew these protoplasmic astrocytes, the prevailing theory was that astrocytes only provided structural support for neurons.  He rejected this idea and instead hypothesized that all the astroglia in the brain formed a sort of gland, which would release substances that affect brain function; it is now known that astroglia do indeed release substances that affect neuronal signaling, including glutamate, GABA, and ATP.  

Having observed the close association of astrocytes with blood vessels and neurons, as well as the fact that all astrocytes appeared to have a prominent appendage, or “foot,” Cajal further hypothesized that astrocytes used this “foot” to stimulate blood vessel dilation.  Astrocyte “feet” do indeed regulate blood vessel diameter, albeit via the release of signaling molecules rather than through physical manipulation, as Cajal imagined.  Astrocyte-evoked changes in blood flow, and thus in oxygenation, constitute the signal that is measured by fMRI, a tool used to image brain activity.

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In this drawing, Cajal details the finer structure of hippocampal dentate granule cell axons, which make mossy fiber synapses onto pyramidal cells in the CA3 region.  The relatively narrow focus of this piece allowed Cajal to depict a wealth of detail in both the neurons themselves and the connections between them.  

The axons of the granule cells in the dentate gyrus delicately contact the proximal dendrites of the giant pyramidal cells in CA3, while the pyramidal cell axons stretch toward the fimbria, the output of the hippocampus, or send collaterals toward CA1.  

Cajal’s major contributions to the field of neuroscience are on display here: the distinct cells, separate yet intertwined, signal the prevailing neuron doctrine, while the arrows signify Cajal’s remarkable intuition regarding the arrangement of the axons and dendrites and the direction of signal transmission according to his law of ‘Dynamic Polarization.’

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Cajal inferred the flow of information between neurons from their structure and relative position.  His ‘Law of Dynamic Polarization’ posits that each neuron is polarized: it has dendrites, through which signals are received, and an axon through which signals are transmitted to the dendrites of the next cells in the pathway.  Thus, simply by observing the morphology and location of neurons in a tissue, he was able to discern the direction of signal transmission.

While this was relatively straightforward in the retina, which receives outside stimuli arrive from a particular direction and must be carried inward, Cajal’s real genius was revealed in the deduction of information flow in a tissue such as the hippocampus, where the sites of input and output were not immediately obvious.  

The hippocampus, named for its resemblance to a seahorse (genus Hippocampus), comprises the Horns of Ammon (Cornu Ammonis, or CA regions), the dentate gyrus, and the subiculum.  It receives signals from various parts of the brain via the entorhinal cortex, and signals flow through the hippocampus in the path shown by Cajal above (dentate gyrus → CA3 → CA1); its output travels through the fornix to the anterior thalamic nuclei and other destinations.  Prior to Cajal’s observations, the fornix was thought to be a source of input to the hippocampus.

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