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Neurohistology is the study of the microstructure and composition of nervous tissue through the use of various staining methods. In most cases, a Hematoxylin and Eosin (H&E) stain is used to stain the nervous tissue (which makes up the nervous system). There are two nervous systems in our body — the central nervous system (CNS) and the peripheral nervous system (PNS). The PNS further divides into the somatic nervous system and the autonomic nervous system. The microstructure of the nervous tissue within and between both major systems are unique as the CNS consists of the brain and spinal cord, while the PNS consists of all other neurons and neuroglia. A neuron is the basic structural and functional unit of the nervous system. Neurons are specialized to receive chemical stimuli then generate and propagate electrical signals. The functions of a neuron are closely related to the structure of the cell. The neurons are made up of a cell body, axon and dendrites. A cell body contains the nucleus of the neuron and it can be stained using a basophilic dye. The stained portions of the cell body are called nissl bodies. Dendrites are specialized to receive nerve signals from receptors or other relating neurons. Axons are the extensions of the cell body which are involved in transduction of the electrical impulses, i.e. the action potential, along its length. When the neuron is excited, the axon transports neurotransmitters (a chemical messenger) and releases them at synapses (a gap between two neurons). A neuron has a long life-span and does not undergo cell division. A sufficient nutritional environment is required for maintaining functional ability throughout life. Additionally, some injured neurons have difficulty recovering after damage. The neuroglia are the surrounding non-neural connective tissues that support the neurons. The CNS has three types of neuroglial cells: astrocytes, oligodendrocytes and microglia. The PNS has one type of neuroglial cell, schwann cells (lemmocytes).

Staining Methods
There are 9 methods used to stain tissue:


 * H&E Staining: H&E staining is a very common staining method. The cytoplasm (a solution that fills the inside of a cell membrane) is acidophilic, so it turns red when stained. The nucleus and its contents are basophilic, so they turn blue.


 * Luxol-fast blue (LFB) staining: In the LFB staining, the myelin sheath (made up of fatty substances and protein) is stained blue.


 * Nissl staining: Using cresyl violet, the Nissl staining method, stains the Nissl bodies purple.


 * Kluver-Barrera (KB) staining: KB staining is a combination of Nissl (using cresyl violet) and LFB staining. The myelin sheath is stained navy blue. It is the most common staining method specific to the nervous system.


 * Bodian silver staining: Uses silver protein to stain neuronal cell bodies (soma) and nerve processes of normal and abnormal structures reddish purple or dark brown. It is often used to detect neurofibrillary tangles, a primary indication of Alzheimer's disease.
 * Holzer staining: Holzer’s crystal violet stains fibrous astrocytic processes blue purple. It is used to detect gliosis, the proliferation of astrocyte fibrous components due to secondary scarring after a nerve damage.
 * Gallyas-Braak (GB) staining: An argyrophilic (affinity to silver) stain that stains abnormal tissues black but leaves normal existing tissues unaffected. It is used to detect pathological structures like tau accumulation.


 * Other various silver stains
 * Bielschowsky and Grocotts's methenamine silver staining: Bielschowsky's silver stain stains nerve fibers and senile plaques in the CNS black. Grocott's methenamine silver stain is used to detect the amyloid components of senile plaques. They are both widely used in the Alzheimer's disease studies.

Brain
The brain is composed of three main parts, the cerebrum, cerebellum and brainstem. The cerebrum, the largest segment of the human brain, is split into two; the left and right hemisphere. The left hemisphere controls language skills, while the right hemisphere deals with self-awareness and spatial information. The outer layer of both hemispheres is called grey matter (cortex) and the inner layer is the white matter. The cerebellum is located at the back of the head, right behind the top of the brainstem. Its main function is to control motor function and muscle coordination. The brainstem connects the cerebellum to the spinal cord, and plays an important role in monitoring consciousness and the respiratory and cardiac (circulatory) systems.

Cerebrum
The cerebrum is covered by pia mater. Pia mater, one of the three protective layers of the meninges, is the innermost protective layer of the brain. In some cerebrum histology slides, the pia mater can be seen. In two different forms of staining, luxol/toluidine blue and eosin stains, six sub-layers are distinguished: I. Outer molecular (plexiform) layer

II. Outer granular layer

III. Outer pyramidal layer

IV. Inner granular layer

V. Inner (ganglionic) pyramidal layer

VI. Multiform cell layer

The cells of the cerebrum are not as organised as the cerebellar cortex.

Cerebellum
Alternatively, the cerebellum only has 3 layers:

I: Outer (molecular) layer

II: Middle (Purkinje cell) layer

III: Inner (granular) layer.

When using H&E staining, Purkinje cells are visible.

The extension of the Purkinje cells through the molecular layer form a dendritic tree. The axons of the Purkinje cell go through the granular layer and deeper into the white matter of the cerebellum. Using a unique silver dye on a cut section that is thick enogh, the dendritic tree and axon of the Purkinje cell can be seen.

Brainstem


The brainstem is made up of 4 parts: the pons, medulla oblongata, diencephalon and midbrain. The medulla oblongata is the lowest part of the brainstem. A well functioning medulla oblongata is critical for survival. It controls the involuntary functions of the heart and lungs.

The medulla oblongata has large neurons. The dark sports within the neurons are nissl substances or bodies. In the image on the left, the larger concentrated purple areas with white space around them, are the neurons in the medulla oblongata. In the same image, the dark dots within the neurons are nissl substances. Nissl substances are made out of endoplasmic reticulum and free ribsosomes within the neurons. The endoplasmic reticulum and ribosomes are organelles (structures that perform specific tasks inside cells).

Spinal Cord


The spinal cord is surrounded by the meninges (a protective layer that covers the brain), in the spinal canal. The spinal canal is a passageway that houses the spinal cord. The spinal cord has 31 different sections, but it is grouped in 5 main segments: cervical, thoracic, lumbar, sacral and coccygeal. The cervical has 8 sections, the thoracic has 12, the lumbar has 5, the sacral has 5 and the coccygeal has 1. The spinal cord is a continuous cord that is made up of different sections.

Aside from different sections, the spinal cord also has gray and white matter, like the brain. A horizontally cut section of the spinal cord in the cervical section, for example, will showcase the H shape or butterfly appearance of the grey matter in the center. Depending on the spinal cord level, the shape of the butterfly is different. The ratio between the grey and white matter increases down the spinal cord (towards the sacral spinal cord region).

In the image on the right, the first drawing in the top left corner is a cervical section, along with the drawing to the right of it. The last drawing in the row is a thoracic section. In the last row, the first drawing is a lumbar section, and on its right is the sacral section. The last two drawing showcase progressive changes in the spinal cord towards the end of the sacral region.The projections of the grey matter facing a person's back are the dorsal horns, the forward facing projections are called the ventral horns. Everything surround the grey matter is comprises of the white matter. The dorsal and ventral horns create 3 different sections of white matter; dorsal (funiculi), lateral (funiculi) and ventral (funiculi). The dorsal and ventral funiculi sandwich the lateral funiculi, so the lateral funiculi is the middle portion of the white matter. All parts of the spinal cord work together to control the voluntary movement of the 4 limbs and receive sensory information from these areas.

Peripheral Neuron and Nerve
The neurons of the PNS are called peripheral neurons. Peripheral neurons are distinct from that of CNS by the followings:


 * 1) Myelination by Schwann cells: Axons are morphologically divided into two types, myelinated and non-myelinated. The nerves of the PNS are myelinated and the myelin of the neurons in the PNS are formed by Schwann cells (lemmocytes). Each Schwann cell myelinated about 100µm of an axon and forms the lipid-rich myelin sheath. The myelin sheath appears as like spokes of a wheel around the axon under the light microscope. It acts as an electrical insulator which allows fast nerve impulse transduction speed along the axon. It can be stained by H&E or Triple-staining. The chemical composition of the myelinated axons in the PNS is also different from that from the CNS.
 * 2) Forms a primary structure called nerve: The peripheral nervous system divides into somatic nervous system and autonomic nervous system. The primary structure of the PNS is a nerve. The axons of the peripheral neurons are bundled together to form fascicles. A fascicle consists of connective tissue, axons, and Schwann cells and it is surrounded by a connective tissue layer called perineurium. Epineurium wraps the entire fascicles and perineurium to form the outermost layer of the peripheral nerve. Such organization helps to create a structured pathway for the neuron function.

Somatic Nervous System




The somatic nervous system is a single neuron system which innervates skeletal muscle to control body movements involving skin, muscle and joints, or receives impulse at somatosensory receptors and transmits it to CNS. It consists of afferent nerves or sensory nerves, and efferent nerves or motor nerves. The sensory neuron is a unipolar neuron where its cell body is at the side. It has a short axon and one long dendron. The motor neuron is a multipolar neuron which has one long axon and many short dendrites. The cell body of the motor neuron is at one end of the neuron length and another end of the axon connects with a skeletal muscle fiber.

Autonomic Nervous Systems
The autonomic nervous system divides into the sympathetic nervous system and the parasympathetic nervous system. The pathway of both system consists of two neurons and a group of neuron cell bodies called ganglion sits in between them. The neuron comes before the ganglion is called pre-ganglionic neuron and the neuron which comes after the ganglion is post-ganglionic neuron. The sympathetic nerves have a short pre-ganglionic neuron and the parasympathetic nerves have a short post-ganglionic neuron.

Astrocytes
Astrocytes are one type of glial cells in the CNS supporting nerve tissues at the blood-brain barrier. Astrocytes have a small cell body and many processes that split into all directions, resembling the shape of a “star”. Two forms of astrocytes are radial and fibrous. Fibrous astrocytes are usually found in the white matter and its process is thin and long with few branches. A large number of filaments are present between the cell body and the processes. Radial astrocytes are highly branched and mainly found in the grey matter. Radial astrocytes have less filaments than fibrous astrocytes.

Astrocytes are involved in the ionic homeostasis, energy metabolism, and synaptic signalling of the brain.



Oligodendrocytes
Oligodendrocytes are one of the glial cells of the CNS. They protect the neurons by producing myelin sheath. The myelin sheath covers the axon to act as an insulator and allow for the signal to be transmitted easily along the axon. The nucleus of the oligodendrocytes are characterised by their round nucleus.

Microglia
Microglia are the non-neuronal cells of the CNS originated from mesoderm, which function as resident macrophages and remove degenerated neurons and foreign substances. Microglia contain branched and thin cytoplasmic processes.

Schwann Cells (Lemmocytes)
Schwann cells are the only glial cells of the PNS. They are also able to myelinate cells, like oligodendrocytes. In addition, they are able to encourage the regeneration of cells after injury. Using H&E staining, the nuclei of the schwann cells will be dark purple.