HistoNCR 2024 - Teórico 3 - 13/8/2024

Understanding White and Gray Matter in the Nervous System

Introduction to Self-Evaluation Questions

• The session begins with a discussion about self-evaluation questions designed to ease participants into the topic, ensuring they are not overwhelmed.

Observations of White Substance

• Discussion on identifying white substance in images, noting that certain areas appear more clearly than others due to ganglion presence.

• Explanation of staining techniques (e.g., Matxin de Osina) used to visualize cellular structures, emphasizing that not all stained fibers can be categorized as white matter.

Distinctions Between Gray and White Matter

• Clarification on the differences between gray matter (neuropil region) and white matter, highlighting their distinct structural characteristics.

• Description of how gray matter appears denser compared to the more aerated structure of white matter, which has identifiable axonal cuts.

Cerebellar Structure Insights

• Examination of molecular layers in both cerebellum and cerebral cortex, noting low neuron density but high neuropil presence for synaptic connections.

• Importance of recognizing structural differences between gray and white substances through specific staining techniques like Clu Barrera.

Identification of Neuronal Structures

• Visual identification of organized gray substance in nuclei versus layered organization in cerebral cortex; introduction to peripheral nervous system components.

• Discussion on deep cerebellar nuclei visibility under microscopy, contrasting it with other regions lacking clear differentiation.

Neuropil Characteristics

• Justification for distinguishing neuropil based on its absence in certain glial cells; emphasis on local connections within the enteric nervous system.

Summary of Class Content

• Recap that while some images show clear distinctions between gray and white substances, others may require deeper analysis using electron microscopy for clarity.

Future Learning Directions

• Acknowledgment that class content was dense; plans to cover synapses and various types of glia in future sessions. Students are encouraged to study recommended chapters thoroughly for comprehensive understanding.

Concept of Synapse

Definition and Types of Synapses

• Synapses are specialized contacts between two cells, with at least one being a neuron. They can be classified as interneuronal synapses (between neurons) or synapses between neurons and effector cells, which may vary in origin and nature.

• These structures serve as specialized communication channels for transmitting information, characterized by specific structural and functional features tailored to their roles.

Neuron Connectivity

• The number of neurons is astronomical; each neuron can form between 1 to 250,000 synapses depending on the cell type. For instance, auditory pathway neurons may establish only a few synapses while Purkinje cells in the cerebellum can create around 250,000.

• Despite the vast number of synapses exceeding that of individual cells, there exists a non-random pattern of connectivity influenced by local circuits and the role each neuron plays within them.

Classification Criteria for Synapses

• Synapses can be classified based on location (e.g., axodendritic or axosomatic), structural components involved in the synaptic contact, and ultra-structural appearance observable via electron microscopy.

• Functionally, synapses can exert excitatory or inhibitory effects on connected cells. This effect primarily depends on the receptors present in the postsynaptic cell rather than solely on the presynaptic cell's characteristics.

Neurotransmitter Effects

• A single neurotransmitter like acetylcholine can have opposing effects (excitatory or inhibitory) depending on the types of receptors it interacts with in different cell types.

• Synapse classification also includes identifying neurotransmitters involved (e.g., glutamatergic or GABAergic), contributing to what is known as a neuron's neurochemical identity.

Types of Synaptic Transmission

• Chemical synapses are predominant in vertebrate nervous systems; they transmit nerve impulses through neurotransmitter release from presynaptic terminals into the synaptic cleft where they interact with postsynaptic elements.

• Neurotransmitters are typically stored in vesicles before release into the synaptic cleft. Some neurotransmitters do not require vesicular storage but will not be discussed here today.

Electrical and Mixed Synapses

• Electrical synapses involve direct cellular contact through gap junctions allowing small molecules like ions and ATP to pass between adjacent cells without neurotransmitter release.

• In electrical transmission, ionic currents propagate directly from one cell to another. Some mixed-type synapses combine chemical and electrical signaling patterns resulting in both rapid and slower cellular responses.

Understanding Chemical Synapses

Structure of Chemical Synapses

• Chemical synapses are characterized by a presynaptic element that releases neurotransmitters, a synaptic cleft, and postsynaptic receptors that bind to these neurotransmitters to affect the receiving neuron.

• The presynaptic element, also known as the terminal button of an axon, can be singular or multiple. Axons typically branch into telodendria, leading to several terminal buttons.

• The axonal terminals contain cytoskeletal elements like microtubules and actin filaments, abundant mitochondria, smooth endoplasmic reticulum, and synaptic vesicles that store neurotransmitters.

Synaptic Cleft Characteristics

• The synaptic cleft is the space between the presynaptic and postsynaptic membranes. It contains dense glycoprotein material and cell adhesion proteins that contribute to its structural integrity.

• Postsynaptically, there can be neurons or other cell types such as muscle cells. Electron microscopy reveals classic images of chemical synapses in the central nervous system.

Visualizing Synapse Structures

• In electron microscopy images of dendrites with two presynaptic elements visible, various structures including mitochondria and synaptic vesicles appear distinctly due to their shapes.

• The presynaptic membrane shows a clear distinction from the postsynaptic membrane across an electron-lucent line representing the synaptic cleft.

Vesicle Types and Synapse Density

• Different shapes of vesicles (round vs. oval) indicate varying types; round vesicles are likely spherical while oval ones may be flattened.

• Observations show that the presynaptic density is thinner than the postsynaptic density; this difference is crucial for identifying asymmetric synapses associated with excitatory functions.

Asymmetrical Synapse Analysis

• An asymmetrical synapse features a thinner presynaptic density compared to a thicker postsynaptic density; this structure often correlates with spherical vesicles indicative of excitatory transmission.

• This structural relationship helps identify functional characteristics of chemical synapses—specifically those classified as excitatory based on their morphology rather than strict rules.

Identifying Neuromuscular Junctions

• A neuromuscular junction is identified by observing regular-sized vesicles associated with membrane densities indicating it’s a presynaptic element linked to skeletal muscle fibers through characteristic cytoskeletal organization.

Neuromuscular Synapse Structure and Function

Overview of the Neuromuscular Junction

• The neuromuscular junction is characterized by a membrane structure located opposite the presynaptic membrane, featuring invaginations that increase postsynaptic density.

• When vesicles fuse and release their contents at active sites into the synaptic cleft, they bind to receptors on the postsynaptic membrane, leading to ion channel openings for sodium influx and potassium efflux.

Mechanism of Action Potential Transmission

• The action potential travels through muscle cell membranes via T-tubules, coupling excitation with contraction through proximity to endoplasmic reticulum cisternae.

• The synapse consists of a linear presynaptic element and an intricate postsynaptic membrane structure.

Types of Synapses in Muscle Fibers

• In skeletal muscle fibers, the neuromuscular junction forms where nerve fibers contact muscle fibers; however, smooth muscle connections differ as they involve more distant structures called en passant boutons.

• En passant boutons are less physically connected to smooth muscle fibers and are surrounded by Schwann cells that facilitate neurotransmitter storage and release.

Presynaptic Structures and Vesicle Dynamics

• A single axon can branch out at its terminal portion, forming multiple presynaptic structures known as terminal buttons.

• The presynaptic element comprises a dense matrix with projections that interact with synaptic vesicles.

Neurotransmitter Release Process

• Synaptic vesicles undergo a cycle similar to previously studied vesicular trafficking processes involving constitutive and regulated pathways.

• An action potential triggers voltage-dependent calcium channels in the presynaptic membrane, leading to vesicle fusion and neurotransmitter release.

Recycling Mechanisms Post-Neurotransmitter Release

• After neurotransmitter release, the presynaptic membrane is recycled via clathrin-coated vesicles which re-form from endosomes.

• Peptide neurotransmitters are synthesized in rough endoplasmic reticulum (RER), processed in Golgi apparatus, then transported along axons to be released at terminals.

Vesicular Transport Proteins

• The recycling process involves proteins like SNARE complexes that mediate vesicle anchoring to target membranes during fusion events.

Neuronal Synapses and Vesicle Dynamics

Characteristics of Presynaptic Structures

• The electron micrograph displays characteristic images of vesicles at varying distances from the presynaptic membrane, with some closely attached and others fused.

• The "Omega" image is described, where fusion occurs between the presynaptic membrane and vesicular membrane, resembling the Greek letter Omega in cross-section.

Neuromuscular Junction Observations

• A neuromuscular junction is shown through cryofracture electron microscopy, highlighting calcium channels and a pool of vesicles associated with the presynaptic membrane.

• Stimulation of nerve fibers reveals active site structures correlating with Omega-shaped structures indicative of vesicle fusion.

Temporal Dynamics of Vesicle Cycling

• Over time, these structures retract from calcium channel regions, leading to the appearance of clathrin-coated vesicles. This illustrates the temporal cycle: formation, anchoring, fusion, and recycling of membranes observed via electron microscopy.

Postsynaptic Density Composition

• Postsynaptic densities primarily consist of various receptors, ion channels, and accessory proteins that create a scaffold for receptor localization while linking postsynaptic elements to the cytoskeleton.

Types of Interneuron Synapses

• Interneuron synapses involve connections between two neurons categorized based on cellular elements in contact. Common types include axosomatic (axon to soma), axodendritic (axon to dendrite), axoaxonic (axon to another axon), and dendrodendritic (dendrites connecting).

Unique Features of Dendrodendritic Synapses

• Dendrodendritic synapses are characterized by reciprocal connections without an axon; both dendrites have synaptic vesicles and postsynaptic densities.

Visual Representation of Synapse Types

• An illustration depicts various synapse types: axosomatic, axodendritic, and axo-spinous. Axodendritic refers to terminal buttons connecting with dendrite trunks while axo-spinous connects with dendritic spines.

Vesicle Morphology Insights

• Vesicles can be spherical or flattened; their morphology may vary due to sample processing but provides insights into synapse type identification.

Differentiation Based on Content

• Electron-dense vesicles like G-type store catecholamines while larger dense-core vesicles contain neuropeptides. Their size helps identify potential synapse types during analysis.

Asymmetrical vs Symmetrical Synapses

Neurotransmission and Synaptic Types

Overview of Neurotransmitter Action

• The microphotograph illustrates the inhibitory action on target points, showing a terminal button with clear central vesicles, which may correspond to catecholamines or neuropeptides based on size.

Diversity of Neurotransmitters

• Multiple neurotransmitters can be associated with a presynaptic button, indicating that neurotransmission involves various neurotransmitters exerting different effects on synaptic targets.

Classification of Synapses

• Synapses are categorized into those with spherical vesicles and asymmetrical densities (Type 1) versus flattened vesicles and symmetrical densities (Type 2), with Type 1 being excitatory.

Clarification on Synapse Types

• A correction will be provided regarding the classification of synapses; Type 1 synapses are asymmetrical while Type 2 are symmetrical.

Characteristics of Grey's Classification

• Grey's classification primarily applies to cortical synapses but is often used broadly across the central nervous system. Type 1 synapses typically have spherical vesicles and are excitatory.

Features of Inhibitory Synapses

• Type 2 synapses possess flattened vesicles and symmetrical densities, generally functioning as inhibitory connections.

Distribution in Neuronal Structures

• Excitatory Type 1 synapses frequently localize at dendritic spine heads, while both types can occur along dendritic trunks and somas, though inhibitory ones are more common in certain areas.

Correlation Between Structure and Function

• Different types of synapses exhibit varying frequencies in specific regions, correlating with circuit-specific functions where each type serves distinct roles.

Detailed Examination of Vesicle Types

• Spherical vesicles indicate fine presynaptic density for excitatory Type 1 synapses compared to symmetrical structures associated with inhibitory connections.

Identifying Synapse Types through Markers

• Specific markers can confirm the neurochemical identity of a given synapse; for instance, examining configurations helps determine whether they are excitatory or inhibitory based on their structure rather than just neurotransmitter nature.

Structural Configurations in Synaptic Connections

• An axodendritic configuration shows how myelinated axons lose their sheath to form terminal buttons connecting to dendrites.

Variations in Axo-Spine Connections

• The presence of irregularly shaped vesicles suggests an axospinous connection that could likely be either excitatory or inhibitory based on structural characteristics.

Complexity in Synaptic Arrangements

• The formation sequence of synapses reflects neural circuit organization: series connections align sequentially while reciprocal ones function bidirectionally.

Differences Between Electrical and Chemical Synapses

Understanding Mixed Synapses and Glial Cells

Mixed Synapses

• Discussion on the existence of mixed synapses, which feature both types of synaptic connections. An electron micrograph illustrates a synapse between two cellular elements, highlighting synaptic vesicles and densities.

• A mixed synapse is defined as a region with synaptic contact that incorporates both electrical and chemical synapses.

Types of Glial Cells

• Overview of glial cells in the central nervous system (CNS), categorized into two main groups: astroglia and microglia. Astrocytes, ependymocytes, and oligodendrocytes share an embryonic origin from the neural tube epithelium.

• Microglia originate from mesodermal cells in the bone marrow, migrating to the CNS around mid-gestation. They are smaller than astrocytes and have distinct functions within the nervous system.

Peripheral Nervous System Glia

• In the peripheral nervous system (PNS), glial cells arise from neural crest cells alongside neurons. Two primary types include satellite cells located in ganglia and Schwann cells associated with axons at synaptic terminals.

Functions of Glial Cells

• Glial cells perform numerous essential functions similar to neurons but are not excitable electrophysiologically. They provide mechanical support, trophic functions, regulate extracellular metabolite levels, protect neurons, form controlled compartments for neuronal function, synthesize myelin sheaths for axons, eliminate dead cells and debris, protect against pathogens, and play roles in tissue degeneration and regeneration.

Macroglia vs Microglia

• The macroglia includes astrocytes (with protoplasmic or fibrous types found in gray or white matter respectively) and oligodendrocytes present in both gray and white matter due to their role in myelination.

• Ependymal cells line ventricles; astrocytes exhibit sun-like branching patterns while microglia display irregular branching with fusiform somas. Oligodendrocytes have fewer extensions but can create complex branching patterns.

Interactions Within Nervous Tissue

• Emphasis on how neuron-glia interactions create an ecosystem that defines nervous tissue properties. Understanding these relationships is crucial for grasping overall brain function.

Astrocytes: Structure and Function

Characteristics of Astrocytes

• Astrocytes are effectively marked using the Golgi silver impregnation technique, revealing protoplasmic astrocytes that resemble pom-poms in appearance.

• The term "pom-pom" is a descriptive adjective for the shape of these cells, which appear fluffy due to their structure.

• Fibrous astrocytes can be identified through immunochemistry for GFAP (Glial Fibrillary Acidic Protein), showcasing longer and less branched processes compared to protoplasmic astrocytes.

Distinctions Between Astrocyte Types

• Fibrous astrocytes exhibit irregularly shaped cell bodies with long, sparsely branched processes; this contrasts with the more compact structure of protoplasmic astrocytes.

• GFAP staining highlights differences in cellular morphology between fibrous and protoplasmic astrocytes, with fibrous types showing dense GFAP presence throughout their extensions.

Functional Roles of Astrocytes

• Protoplasmic astrocytes have finer extensions that are less visible under GFAP staining, making them harder to identify in gray matter compared to fibrous astrocytes in white matter.

• Generally, astrocyte nuclei are oval or rounded with prominent chromatin patterns; they contain minimal granules but abundant organelles.

Historical Perspective on Astrocyte Function

• Santiago Ramón y Cajal referred to astrocytes as the "second element," emphasizing their structural and functional importance beyond mere support for neurons.

• Astrocytes provide not just structural scaffolding but also create an environment conducive to neuronal activity, acting as a framework where neural interactions occur.

Structural Support Provided by Astrocytes

• Rich in intermediate filaments, astrocytes contribute significantly to mechanical resistance within nervous tissue while forming a supportive network for neuronal processes.

• Their terminal expansions contact various components of nervous tissue, including blood vessels, establishing boundaries between extracellular spaces and vascular walls.

Interaction with Neural Structures

• Astroctyes form glia limitans layers that delineate nervous tissue from surrounding structures like meninges; they play crucial roles at synaptic interfaces.

• Electron microscopy reveals how perivascular feet of astroctyes envelop capillaries, creating interfaces essential for maintaining homeostasis within the brain's microenvironment.

Variations Among Different Types of Astroctyes

• There are distinct types of astroctyes such as radial glia involved during development; others like Bergmann glia in the cerebellum serve specific functions related to their anatomical locations.

Astrocytes and Their Role in Synaptic Regulation

Functions of Protoplasmic Astrocytes

• Protoplasmic astrocytes play a crucial role in synaptic regulation, primarily located in gray matter where synapses occur, unlike white matter which lacks synapses.

• In white matter, astrocytes are associated with the nodes of Ranvier due to the abundance of myelinated fibers, highlighting their specific functions in different brain regions.

Structural Organization and Communication

• Astrocytic processes are intricately linked to synapses, demonstrating their pervasive presence throughout nervous tissue while maintaining organized territories that overlap minimally with neighboring astrocytes.

• These astrocytic territories contribute to the spatial organization of neural tissue and establish gap junction contacts with adjacent astrocytes, forming a network that facilitates intercellular communication.

Calcium Waves and Synaptic Interaction

• Astrocytes can generate calcium waves that propagate over long distances within neural tissue, influencing local environments and interactions with synapses.

• The connections between astrocytic processes and dendrites illustrate the historical recognition by neurologists of astrocyte involvement in synaptic function.

Evolution of Synapse Models

• Traditional models depicted synapses as bipartite structures (pre-synaptic and post-synaptic), but recent findings have led to a revised model known as the "tetrapartite" synapse.

• This new model includes pre-synaptic elements, post-synaptic elements, and associated astrocytic processes that actively participate in neurotransmitter uptake and modulation of synaptic transmission.

Microglia: Structure and Function

• Microglia are branched cells found in both gray and white matter; they possess an irregular oval nucleus with less cytoplasm compared to astrocytes.

• Their ramified extensions exhibit an undefined branching pattern, contrasting sharply with neuron morphology. Specific immunohistochemical techniques help visualize microglial structure effectively.

Dynamic Behavior of Microglia

• Live imaging reveals microglial behavior; although their cell bodies remain relatively stationary, their processes continuously survey the extracellular environment for interaction opportunities.

• Observations show microglial processes retracting or extending dynamically as they interact with neighboring cells' surfaces.

Microscopy and Cellular Dynamics

The Nature of Microscopy in Cellular Studies

• Classical optical microscopy presents limitations as it fails to capture the dynamic nature of cellular elements like microglia, which are constantly in motion.

• Synaptic changes lead to retraction of dendrites, with microglia playing a role in phagocytosing terminal ends during these processes.

Functional States of Microglia

• Microglia can exist in various functional states; the "quiescent" state acts as a sentinel, continuously sensing the environment through its extensions.

• An "ameboid" form appears during embryonic development and injury responses, while activated microglia exhibit larger somas and more pronounced extensions.

Response to Infections

• Activated microglia respond to pathogens by expressing major histocompatibility complex (MHC) proteins, crucial for antigen presentation and pathogen recognition.

Oligodendrocytes: Structure and Function

Characteristics of Oligodendrocytes

• Oligodendrocytes are small cells with few branches found in both gray matter (alongside neuronal somas) and white matter (in rows).

• They form myelin sheaths around axons, characterized by low cytoplasm density but high electron density.

Myelination Process

• Each oligodendrocyte can produce multiple segments of myelin sheath—up to 50 segments—around different axons despite having few initial extensions.

• This ability allows oligodendrocytes to create internodal segments between nodes of Ranvier effectively.

Comparative Analysis: Oligodendrocytes vs. Schwann Cells

Differences in Myelination

• Unlike oligodendrocytes that can myelinate multiple axon segments, Schwann cells in the peripheral nervous system only myelinate one segment each.

Structural Insights from Electron Microscopy

• Observations reveal complex structures resembling candelabras formed by oligodendrocyte extensions creating internodal segments within white matter.

Visualizing Myelin Sheath Formation

Electron Microscopy Findings

• High-resolution images show oligodendrocytes forming successive layers of myelin around adjacent fibers, highlighting their intricate structure.

Internal Structures of Myelin Sheaths

• The internal mesaxon is visible where cytoplasm meets the first layer of the myelin sheath; however, external mesaxon visibility is limited due to space constraints within central nervous system sections.

Complexity of Oligodendrocyte Extensions

Ramification Patterns

• Despite appearing simple at first glance, oligodendrocyte branches can become highly complex when visualized under scanning electron microscopy.

Association with Peripheral Nervous System Cells

Understanding the Structure of Myelinated and Unmyelinated Nerve Fibers

The Role of Schwann Cells and Axons

• Each segment of a nerve fiber is formed by a Schwann cell, with unmyelinated fibers not being naked but surrounded by cells that present longitudinal invaginations where the axon is embedded.

• The term "mesaxon" refers to the contact points between the edges of the axonal prolongation and the surrounding structures, highlighting its structural significance.

Extracellular Matrix in Nervous Tissue

• The extracellular matrix (ECM) has often been overlooked; it consists of cellular elements and matrix components that organize nervous tissue, which appears densely packed in histological images.

• Historical techniques for fixing samples can create artifacts that misrepresent the actual structure, leading to misconceptions about space within neural tissues.

Artifacts in Histological Preparation

• Fixation methods can cause significant volume reduction in tissue samples, primarily due to collapse of extracellular spaces. This highlights discrepancies between fresh and fixed brain tissue.

• Properly processed nervous tissue shows that extracellular space exists and plays a crucial role in cellular communication through signaling molecules.

Properties and Functions of Extracellular Space

• Research into how this extracellular space functions as a drainage system akin to lymphatic systems has gained traction, emphasizing its importance beyond mere structural support.

• Recent studies have begun to reveal ECM's physiological roles within nervous tissue, indicating it contributes significantly to overall functionality.

Components of Extracellular Matrix

• ECM is found throughout central nervous system tissues, including basal membranes around capillaries and nodes. It comprises both diffuse components within neuropil and structured networks around neurons known as perineuronal nets.

• Perineuronal nets consist of various molecular components like hyaluronic acid and proteoglycans, forming a complex network essential for neuronal function.

Visualization Techniques for Extracellular Matrix

• Different staining techniques using lectins—plant-derived proteins that bind specifically to sugars—can help visualize ECM components effectively.

• These techniques allow researchers to observe cellular contours similar to Golgi staining but are distinct due to their specific binding properties related to glycosaminoglycans.

Understanding Perineuronal Networks and Their Role in Neural Circuits

Overview of Perineuronal Networks

• The use of chromogen immunofluorescence reveals a network-like structure known as perineuronal networks, characterized by their porous appearance.

• In the cerebral cortex, a dark homogeneous area corresponds to diffuse components interspersed among neuronal processes observed under electron microscopy.

Visualization Techniques

• Fluorescence microscopy highlights neuronal nuclei in red and perineuronal networks in green, indicating that these networks are primarily found around inhibitory GABAergic interneurons.

• Chemical composition varies across different regions of the matrix, affecting both cellular properties and the diffuse component's characteristics.

Synaptic Structures and Matrix Interaction

• Immunofluorescence illustrates presynaptic terminals (glutamatergic), showing how synapses localize within the holes of perineuronal networks, stabilizing them.

• The relationship between synapses and the extracellular matrix is crucial; it acts as a stabilizer for circuit configurations.

Evolution of Synaptic Models

• Historical models have evolved from bipartite to tripartite synapse structures, with current models proposing a tetrapartite organization involving the extracellular matrix.

Importance of Extracellular Matrix

• The extracellular matrix not only provides structural support but also influences neuronal properties and local circuits significantly.