Wheater's (6th ed.), Ch 20, Central Nervous System

Stevens and Lowe's (4th ed.), Ch 16, Nervous System

Overview:

1. Spinal Cord
2. Neurons
3. Glia
   1. ependymal cells / choroid plexus
   2. oligodendrocytes
   3. astrocytes
4. Cerebellum
5. Cerebrum (neocortex)
6. Hippocampal formation (archicortex)

Slide Descriptions

I. Spinal Cord
a. Slide 65-2 (lumbar spinal cord, H&E) [DigitalScope]
b. Slide 66a (thoracic spinal cord, luxol blue & cresyl violet) [DigitalScope]

Review the organization of the spinal cord using an anatomy atlas, then examine the cross section of the lumbar spinal cord in slide #65-2.  At low magnification, differentiate inner gray matter from outer white matter and identify dorsal and ventral horns of the gray matter. In these slides, dorsal happens to be "up," but you should be able to tell dorsal and ventral horns based on morphology and the cells present rather than the orientation. The perikarya of large somatic motor neurons [example] located in the ventral horn of the cord innervate the skeletal muscles of the limbs and trunk of the body (soma="body" in Greek, hence "somatic").

Recall during development, newly formed neurons proliferate adjacent to the canal to form a mantle layer, which becomes the gray matter in the central region of the mature spinal cord. This gray matter occupies a butterfly-shaped cross-section in this mature spinal cord.  In the gray matter, examine the large motor neuron cell bodies carefully.  Depending on how the cells are cut, you may see the nucleus, nucleolus, and axon hillock, as well as the Nissl bodies (rough ER) filling the cytoplasm.  Note also the smaller nuclei of neuroglial cells.  Neuropil refers to the regions in gray matter that lie between cell bodies, devoid of nuclei but complexly crowded with neuronal cell processes and synapses. 

The neurons of the mantle layer produce processes that grow outward to form the fiber tracts of white matter, which carry parallel myelinated axons for long-range interneuronal contacts in CNS.  The white matter fills most of this cross-section external to the central butterfly-shaped region of gray matter.  This marginal zone is white primarily because of the abundance of myelinated axons, which you can observe in your sections, although the axons are very faint and therefore difficult to see.

Slide 66a shows a section of thoracic spinal cord. In addition to the dorsal and ventral horns, two structures especially obvious in the thoracic cord are the dorsal nucleus of Clarke and the lateral horn. The dorsal nucleus of Clarke [example] is in the dorsal horn and contains relatively large, multipolar neurons that receive proprioceptive information from dorsal root ganglion cells innervating muscle spindles in the trunk and lower limb. The cells of Clarke's nucleus then relay this information via axonal projections that extend all the way up into the cerebellum (hence the reason why the cells are so large) where it is processed to allow for coordinated movement. The lateral horn [example] contains relatively large, multipolar visceral motor neurons of the intermediolateral cell column that extends from levels T1 through L2 of the spinal cord. The cells here are preganglionic sympathetic motor neurons whose axons terminate in either sympathetic chain ganglia or the "visceral" (or "pre-aortic") ganglia associated with the major branches of the abdominal aorta (e.g. celiac, aorticorenal, and superior/inferior mesenteric ganglia). Note that sacral levels of the cord (levels S2-4) also contain visceral motor neurons in the lateral horn, but these are parasympathetic motor neurons.

Many neurons in the spinal cord may appear shrunken and surrounded by an empty space due to poor fixation.  Cells that are well preserved show features characteristic of most neurons: large cell body, large pale nucleus, Nissl substance, and cell processes (most of which are dendrites). The delicate meshwork of dendritic processes and nerve fibers (axons) lying between cells in the gray matter is called the neuropil. The white matter contains nerve fibers (axons) entering and exiting the gray matter, and traveling up and down the spinal cord, linking it to the brain. Another feature commonly found in nervous tissue are starchlike granules known as "corpora amylacea" [example] (amylon = starch, Greek) which are aggregates of dead cells and/or proteinaceous secretions that may be found in either white or gray matter. These granules are of little pathological significance, but they generally increase with age.

Nervous tissue contains two basic categories of cells: neurons and support cells (glia). Both neurons and glia have fine processes projecting from the cell body, which generally cannot be resolved in the light microscope without special staining techniques. Astrocytes in the CNS provide metabolic support for neurons and play an important role in maintenance of the blood-brain barrier whereas oligodendrocytes (another type of glial cell) are responsible for myelination of CNS axons. Recall that Schwann cells are the glial cells responsible for myelination in the peripheral nervous system. Myelin is lipid-rich, and on gross inspection appears white. Thus, in the 'white matter' of the brain and spinal cord, myelinated axons are the predominant neuronal component whereas most of the the nuclei that you see in white matter are primarily of glial cells. The ‘gray matter’ contains relatively more neuronal and glial perikarya as well as non-myelinated (e.g. dendritic) processes. The other major glial cell type you should know about are microglia which are small cells derived from blood monocytes. They are considered part of the mononuclear phagocytic system and will prolifereate and become actively phagocytic in regions of injury and/or inflammation.

The tiny central canal recalls the origin of the nervous system as an infolding which closes off to form the neural tube.  Locate the central canal in the middle of your section and examine the lining layer of ependymal cells.  Compare the ependymal nuclei with the neuronal nuclei you observed in previous slides.  Ependymal cells also line the ventricles of the brain that are continuous with the central canal of the spinal cord.

II. Neurons
Slide 0302_O (cat): Lumbar spinal cord, c.s., TB [DigitalScope]

Neurons [example] are typically characterized by a large cell body or perikaryon containing a large, pale (active, euchromatic) nucleus with a prominent nucleolus.  Scattered in the cytoplasm are the characteristic clusters of ribosomes and rough ER termed Nissl bodies  or Nissl substance. One or more cell processes may also be seen emerging from the neuronal perikaryon. The dendrites receive neural input from other neurons via synapses (or they are specialized to receive sensory stimuli), and they transmit neural information toward the perikaryon. A single axon (often called a nerve fiber) leaves the perikaryon and transmits neural signals to other neurons or to the effector organ (e.g., skeletal muscles) via synapses. The axon may be identifiable by the notable ABSENCE of Nissl substance at the axon hillock; however, this is not always so easy to see by light microscopy.

For this section, you should also

• determine the dorsal and ventral aspects of the cord
• differentiate gray matter from white matter
• distinguish dorsal and ventral horns
• note the central canal lined by ependymal cells

III. Glial Cells

A. Ependymal cells / Choroid Plexus / Oligodendrocytes
NP004N hippocampal region, coronal section, luxol blue [DigitalScope]

Many types of glial cells require special histological stains and can’t be unambiguously identified in regular histological slides.  Ependymal cells, which are (low) columnar epithelial cells lining the ventricles of the brain the central canal of the spinal cord, however, are rather easy to discern.  In this section, lining ependymal cells [example] can be seen all along the interface of the gray matter and the ventricles. Also present in this slide is the choroid plexus [example], which is the source of CSF. Here, specialized ependymal cells actively transport ions into the ventricular space (and water follows). This process relies on membrane-associated Na/K ATPases; thus the cells are quite eosinophilic due to the highe concentration of membrane and mitochondria. The connective tissue immediately below the choroidal cells is also very richly vascularized. 

 

B. Oligodendrocytes are myelinating cells of the CNS and are best found by looking in the white matter tracts within the CNS or in the immediate vicinity of neurons in the cortex. Recall that the white matter tracts are found in the PERIPHERY of the spinal cord, whereas in the cerebrum and cerebellum are organized such that the gray matter is in an outer cortex and the white matter is deeper. Morphologically, oligodendrocytes have round nuclei surrounded by a halo of clear cytoplasm, giving them a bit of "fried egg" appearance. Slide NP004N has been stained with luxol blue, which preferentially stains white matter and can be seen as the dark purple areas beneath the cortex. Most of the nuclei seen within the white matter tracts most likely belong to oligodendrocytes [example], although there are certainly quite a few astrocytes mixed in as well, so the only way to really know for sure is with immunostaining with antibodies specific to oligodendrocytes.

 

C. Astrocytes
Slide WestU-C_001 astrocytes, Gold-staining [DigitalScope]

The links above should open to a view of a lighter stained area of the slide where you can look for typical star-shaped cells, which represent astrocytes.  Many of these astrocytes send out processes that contact and wrap around nearby capillaries, which are also clearly recognizable as tube-shaped segments [example].

IV. Cerebellum
a. Slide 0085_C (cerebellum, cat, Luxol fast blue - Cresyl violet) [DigitalScope]

These slides show a section of the cerebellum in which you can appreciate its very highly folded surface. The gray matter of the cerebellar cortex is organized into an outer molecular layer containing basket and stellate cells (not distinguishable by routine light microscopy) as well as axons of granule cells found in the deeper, highly cellular granule cell layer.  Most of the nuclei visible in the granular layer belong to very small neurons called granule cells, which participate in the extensive intercommunication involved in the cerebellum’s role in balance and coordination. Deeper to the granule cell layer is the white matter of the cerebellum, which contains nerve fibers, neuroglial cells (many of which are oligodendrocytes), and small blood vessels but no neuronal cell bodies.

Examine the boundary between molecular and granule cell layers.  Here you will see the prominent Purkinje neuron cell bodies.  You will not be able to discern the amazing dendritic tree that extends from the Purkinje cell bodies into the molecular layer, nor will you be able to see their axons, which extend down through the granular layer into white matter tracts of the cerebellum.

Deep within the white matter of the cerebellum (at the bottom of the slide), you may notice a collection of large neuronal cell bodies [example], which is likely part of one the so-called "deep cerebellar nuclei" (e.g. the fastigial, globose, emboliform, and dentate nuclei) that we will study in more detail later in the course. 

V. Cerebral Cortex

a. Slide 76 (cerebrum, luxol blue/cresyl violet) [DigitalScope]
Slide 76b (cerebrum, toluidine blue & eosin) [DigitalScope]
  
Unlike the highly organized cerebellar cortex, the cerebral cortex appears to be less well-organized when viewed with the light microscope.  Nonetheless, it is loosely stratified into layers containing scattered nuclei of both neurons and glial cells.  Examine the layered organization of the cerebral cortex using slide 76 stained with luxol blue/cresyl violet [ORIENTATION] (which stains white matter tracts and cell bodies).  Typically one or more sulci (infoldings) will extend inward from one edge of the section.  Examine the gray matter on each side of the sulcus using first low and then high power.  Neurons of the cerebral cortex are of varying shapes and sizes, but the most obvious are pyramidal cells.  As the name implies, the cell body is shaped somewhat like a pyramid, with a large, branching dendrite extending from the apex of the pyramid toward the cortical surface, and with an axon extending downward from the base of the pyramid.  In addition to pyramidal cells, other nuclei seen in these sections may belong to other neurons or to glial cells also present in the cortex.  You may be able to see subtle differences in the distribution of cell types in rather loosely demarcated layers. There are 6 classically recognized layers of the cortex:

1. Outer plexiform (molecular) layer: sparse neurons and glia
2. Outer granular layer: small pyramidal and stellate neurons
3. Outer pyramidal layer: moderate sized pyramidal neurons [example]
4. Inner granular layer: densely packed stellate neurons (usually the numerous processes aren’t visible, but there are lots of nuclei reflecting the cell density)
5. Ganglionic or inner pyramidal layer: large pyramidal neurons [example]
6. Multiform cell layer: mixture of small pyramidal and stellate neurons

Throughout each of these layers are a large variety of inhibitory interneurons that vary in size and morphology. The vast majority of these interneurons are small with cell bodies that are typically smaller than the pyramidal neurons in layer V and axons that only project locally within a few millimeters of their origins in the cerebral cortex.

Pyramidal cells in layers III and V tend to be larger because their axons contribute to efferent projections that extend to other regions of the CNS –pyramidal neurons in layer V of motor cortices send projections all the way down to motor neurons in the spinal cord! (See below.) As you browse through the cerebral cortex in slide 76 and 76b, especially at high magnification, keep in mind that roughly half of the cells that you see are neurons and the other half are glia (and vascular endothelial) cells. Of those that are neurons, roughly half are excitatory (pyramidal, stellate and semilunar neurons) and the other half are inhibitory interneurons.

Deep to the gray matter of the cerebral cortex is the white matter that conveys myelinated fibers between different parts of the cortex and other regions of the CNS. Be sure you identify the white matter in both luxol blue [example] and TB&E-stained [example] sections, as it will appear differently in these two stains. Review the organization of gray and white matter in cerebral cortex vs. spinal cord.(See below.) As you browse through the cerebral cortex in slide 76 and 76b, especially at high magnification, keep in mind that roughly half of the cells that you see are neurons and the other half are glia (and vascular endothelial) cells. Of those that are neurons, roughly half are excitatory (pyramidal, stellate and semilunar neurons) and the other half are inhibitory interneurons.

Deep to the gray matter of the cerebral cortex is the white matter that conveys myelinated fibers between different parts of the cortex and other regions of the CNS. Be sure you identify the white matter in both luxol blue and TB&E-stained sections, as it will appear differently in these two stains. Review the organization of gray and white matter in cerebral cortex vs. spinal cord.

 

c. Slide 303 (human paracentral lobule, thionine stain for Nissl substance) [DigitalScope]

Now that you are oriented to the general presentation of “neocortex” (i.e., newer in evolutionary terms and typically six-layered), inspect slide 303, which was obtained from the paracentral lobule in a parasagittal plane of section. This section is stained with thionine, which highlights the presence of Nissl substance. To orient you to this section, dorsal is toward the top of the image and anterior is toward the left. Near the middle-top of this section, the cortex folds into a sulcus that extends in the ventral and ventral-posterior direction; this is the central sulcus. Thus, the precentral and postcentral gyri are fusing with the paracentral lobule in this section, with the precentral gyrus on the anterior side of the central sulcus and the postcentral gyrus on the posterior side. Look careful at these two cortical gyri and notice their differences. Here are some observations to consider:

• the cortex of the precentral (motor) gyrus is about 50% thicker than the cortex of the postcentral (sensory) gyrus
• the postcentral gyrus appears much more typical, in terms of six recognizable neocortical layers, compared to the precentral gyrus in which the layers are much more difficult to recognize
• the density of cells (neurons and glia) is much greater in the postcentral gyrus compared to the precentral gyrus
• there is more neuropil (unstained space between cell bodies in a Nissl stain) in the precentral gyrus compared to the postcentral gyrus
• the cortex of the postcentral gyrus features a prominent inner granular layer (layer IV), while this layer is very difficult to recognize in the cortex of the precentral gyrus
• layer V of the cortex in the anterior bank of the central sulcus (motor cortex) features the largest neurons found anywhere in the human brain; these large, darkly staining pyramidal neurons are called “Betz cells” [example]

Knowing, as you now do, that the precentral gyrus is the motor cortex and the postcentral gyrus is the primary somatic sensory cortex, why do you think such differences in the cellular architecture (i.e., “cytoarchitecture”) of the cortex exist? Do you think all functional divisions of the cerebral cortex have such recognizable cytoarchitectonic features?

To apply some of what you might be thinking in response to these questions, interrogate the cytoarchitectonic features of the cortex as it forms the precentral and postcentral gyri and folds into the central sulcus. Consider where you would place the boundaries between what we now call Brodmann’s Area (BA) 4 (the primary motor cortex) and Brodmann’s Area 3 (the primary somatic sensory cortex). Similarly, consider where you would recognize a transition somewhere in the crown of the precentral gyrus between BA 4 and the adjacent Brodmann’s Area, BA 6 (the premotor cortex). Finally, do the same for the posterior border of BA 3 where it transitions to BA 1 (yet another division of the primary somatic sensory cortex) in the crown of the postcentral gyrus.

 

d. Slide 304 (human occipital lobe, thionine stain for Nissl substance) [DigitalScope]
e. Slide 305 (human occipital lobe, Gallyas silver stain for myelin) [DigitalScope]

Slides 304 and 305 are adjacent histological sections obtained from a human occipital lobe in a plane very near a coronal plane, but just oblique enough to be roughly orthogonal to the calcarine sulcus. Slide 304 is a thionine stain, which highlights the presence of Nissl substance, and slide 305 is a Gallyas silver stain, which produces a medium brown stain in myelin. In both slides (since these sections are contiguous), the medial face of the hemisphere is toward the bottom and the lateral convexity of the occipital lobe is toward the top. Thus, the calcarine sulcus is the major sulcus formed by the infolding of the cerebral cortex on the medial face of occipital lobe (from the bottom of the slide). The inferior bank of the calcarine sulcus is the lingual gyrus (bottom left in this section) and the superior bank of sulcus is the cuneus gyrus (bottom center in this section). Together, these banks of the calcarine sulcus harbor the primary visual cortex, Brodmann’s Area 17. BA 17 is also known as the “striate cortex” because of a prominent “stria” (stripe or band) of myelinated fibers in layer IV of the neocortex in BA 17 [example]. This myelinated band is named the “stria of Gennari” after the then medical student, Francesco Gennari, who first described it in his 1782 text. This feature of BA 17 is easy to recognize in slide 305. Notice how abruptly this stia terminates in the lingual and cuneus gyri. Indeed, this boundary makes BA 17 the most distinctly bordered and most easily discerned cytoarchitectonic division of the entire cerebral cortex. It is even possible to see the stria of Gennari with the unaided eye in a freshly cut occipital lobe -- the only cytoarchitectonic feature that is visible without a microscope.

Now, compare slides 304 and 305 and see if you can determine how the stria of Gennari fits into cortical layer IV in the Nissl stain. As you attempt to solve this challenge, note the complexity of cortical layer IV as seen in the Nissl stain in BA 17. Indeed, you may find it challenging to recognize the six conventional layers of the neocortex, as described above. You may very well believe that there are at least NINE layers in BA17! This is because of the complexity of cortical layer IV, which is sublaminated into layers IVA, IVB (where the stria of Gennari lies), and a sublamina C that is even further sublaminated into alpha and beta sublaminae. Why do you think this is so? What might be the functional significance of so many sublaminae in cortical layer IV within BA17?

Lastly, as you did for BA 4 and BA 3, see if you can mark the boundaries of BA 17 in both the Nissl and myelin preparations. You may be surprised to know that the vertical meridian (the vertical seam that divides one half of the visual field from the other) is represented in the primary visual cortex precisely along this boundary. Furthermore, across this boundary the mapping of visual space actually reverses (more on “visuotopy” when we study central visual processing later in the course).

VI. Hippocampal Region
NP004N hippocampal region, coronal section, luxol blue [DigitalScope]

This coronal section includes the hippocampus (hippocampus = sea horse), dentate gyrus, and adjacent temporal lobe gyrus (entorhinal cortex). Above the temporal (ventral or inferior) horn of the lateral ventricle the lateral geniculate nucleus is present. Lateral to this structure is the tail of the caudate. The medial surface of the section is the posterior portion of the thalamus and a small portion of the cerebral peduncle. Look at the margins of the ventricle at higher magnification and note that it is entirely lined by ependymal cells. Just medial (to the right) of the tail of the caudate, note the choroid plexus [example], which consists of highly convoluted and vascularized villi covered by ependymal cells which are specialized for the production of cerebrospinal fluid, or CSF.

The hippocampus and dentate gyrus function in what is known as the "limbic system" to integrate inputs from many parts of the nervous system into complicated behaviors such as learning, memory, and social interaction beyond the scope of what can be described here. For now, focus just on the morphology of these regions and observe the presence of three distinct layers rather than the six layers found in the cerebral cortex (evolutionarily speaking, the three-layered organization is considered to be "older," so this type of cortex is also known as "archicortex"). In the hippocampus [ORIENTATION], observe:

• ("1" in the orientation figure) a polymorphic layer containing many nerve fibers and small cell bodies of interneurons,
• ("2" in the orientation figure) a middle pyramidal cell layer containing hippocampal pyramidal cells [example], and
• ("3" in the orientation figure) a molecular layer containing dendrites of the pyramidal cells.

In the dentate gyrus [ORIENTATION], observe:

• ("4" in the orientation figure) a polymorphic layer containing nerve fibers (known as "mossy fibers") and cell bodies of interneurons,
• ("5" in the orientation figure) a middle granule cell layer containing the round, neuronal cell bodies of dentate granule cells [example], and
• ("6" in the orientation figure) a molecular layer containing dendrites of the granule cells.

The "hilus" is the region where the head of hippocampus abuts the dentate gyrus. The multipolar neurons in this area are known as "mossy cells" [example] and they primarily receive input from mossy fibers of the granule cells of the dentate gyrus and then relay those signals back to other cells in the dentate. In terms of clinical significance, the pyramidal cells of the hippocampus are particuarly vulnerable to damage in severe circulatory failure and by anoxia or persistent severe seizures. These neurons and their processes are also among those that are most severely impacted in Alzheimer’s disease, which helps explain why short-term memory loss is one of the cardinal symptoms in dementia. You may see small calcific bodies in part of the hippocampus, which occur as a normal part of the aging process. Calcific bodies are also present in the choroid plexus, another common site of accumulation as the years pass.

On a more positive note, the cells and circuits of the hippocampus are among the most plastic (modifiable) of all the neuronal circuits in the brain owing to the high concentration of NMDA receptors for glutamate expressed by hippocampal pyramidal neurons. Indeed, because of the massed effects of synaptic and circuit plasticity, it is possible to track changes in the size of the hippocampus by MRI in living people as a radiographic marker of health, wellness, injury or disease. In short, just about all variations in human behavior that are considered healthy or adaptive seem to promote exuberant plasticity in the hippocampus and its physical enlargement. The converse is true: just about anything deemed by biomedical or social science to be unhealthy or maladaptive seems to promote destructive plasticity and reduction in hippocampal volume.