BBB-Matrona Chachua

The three main interfaces are:

1. Blood-Brain Barrier (BBB) - Comprises the largest area of blood-brain contact with specialized endothelial cells forming tight junctions.
2. Blood-Cerebral Spinal Fluid Barrier (BCSFB) - Formed by fenestrated endothelial cells at the choroid plexuses and tight junctions of epithelial cells.
3. Arachnoid-Dura Barrier (ADB) - Comprises arachnoid cells underlying the dura mater.

The BCSFB is formed by fenestrated endothelial cells that are leaky, allowing for some transport, while the BBB is formed by specialized endothelial cells with tight junctions that restrict transport more stringently.

Tight junctions in both the BBB and BCSFB restrict the passage of molecules between cells, thereby maintaining the integrity of the barriers and protecting the CNS from unwanted substances.

The primary role of the BBB is to provide a barrier function for the brain by restricting the permeability of brain capillaries, particularly to hydrophilic (water-soluble) molecules, thus protecting the brain from potentially harmful substances while allowing essential nutrients to pass through.

The surface area of the BBB is estimated to be 5000 times larger than that of the BCSFB, making it the largest barrier in the brain.

Tight junctions (TJ), also known as zonulae occludens, are connections between neighboring brain endothelial cells (BECs) that reduce the paracellular movement of molecules, thereby creating a physical barrier that contributes to the integrity of the BBB.

Specialized brain endothelial cells (BECs) are crucial components of the BBB as they line the brain capillaries, form tight junctions, and create metabolic and transport barriers, allowing selective access of essential nutrients while restricting harmful substances.

The vascularization of the brain is extensive, with estimates suggesting that each neuron is perfused by its own blood vessel, highlighting the importance of the BBB in maintaining neuronal health and function.

The choroid plexuses produce cerebral spinal fluid (CSF) with a turnover rate of around four times every 24 hours.

The endothelial cells at the choroid plexus are fenestrated, which is different from the non-fenestrated endothelial cells found in most of the brain.

The BCSFB barrier is constituted by tight junctions (TJs) formed by epithelial cells surrounding the fenestrated endothelial cells at the choroid plexus.

The main components of the BCSFB include:

1. Fenestrated capillary endothelial cells
2. Choroid plexus epithelial cells
3. Cerebral spinal fluid (CSF)
4. Tight junctions
5. Ependyma
6. Extracellular fluid (ECF)

Circumventricular organs (CVOs) are specialized regions in the CNS where blood-brain endothelial cells (BECs) are fenestrated. They are located in the ventricles and primarily serve two functions:

1. Neuropeptide secretion (e.g., pineal gland)
2. Hormone/neurotransmitter detection (e.g., subfornical organ)

At circumventricular organs, specialized ependymal cells or tanycytes are linked by tight junctions (TJs) to act as a barrier.

In the blood-CSF barrier (BCSFB), tight junctions connect the choroid plexus epithelial cells and ependyma, helping to regulate the movement of substances between the blood and cerebrospinal fluid (CSF).

The BBB regulates ionic gradients by maintaining specific intra- and extracellular concentrations of ions, particularly K+ and Na+. This regulation is crucial for efficient neuronal firing and neurotransmitter release, as these ionic concentrations must remain within a narrow range.

The BBB controls the exchange by preventing the entry of potentially neurotoxic elements from blood plasma into the brain. For instance, it restricts the passage of high levels of excitatory neurotransmitters like glutamate and high-molecular weight proteins such as albumin, which can induce neuronal apoptosis.

The BBB allows nutrient exchange by facilitating the transport of essential nutrients and signaling molecules, such as glucose, into the brain. This is crucial as the brain consumes 20% of the body's oxygen and glucose, and it relies on transporters like glucose transporter-1 (GLUT1) to access these nutrients due to the exclusion of many hydrophilic molecules by the BBB.

The BBB and choroid plexus control the composition of ECF by producing cerebrospinal fluid (CSF) and regulating the interstitial fluid (ISF). The CSF cushions the brain, reduces its weight, and provides drainage for ECF. The BBB also facilitates slow ISF drainage through osmotic and ionic gradients, ensuring proper fluid balance and composition.

The neurovascular unit describes the interactions between:

1. Brain Endothelial Cells (BECs) - Lining the capillaries and interconnected by tight junctions (TJ).
2. Pericytes - Small connective tissue cells attached to the abluminal surface of BECs, containing smooth muscle myosin.
3. Astrocytes - Glial cells with end feet that surround the basal lamina.
4. Neurons - Nerve cells that interact with the other components.
5. Neighboring Glia - Other supportive cells in the brain.

The basement membrane (BM) is composed of various extracellular matrix proteins, including:

• Structural proteins: Collagen type IV
• Adhesion proteins: Laminin, Fibronectin
• Heparan sulfate proteoglycans: Perlecan, Agrin

These components play a crucial role in the integrity and function of the BBB.

Integrins are the major class of receptors expressed by all cell types in the neurovascular unit. They exist in different subclasses that allow binding to various components of the basement membrane, such as collagen and laminin, facilitating cell adhesion and communication within the BBB.

Matrix metalloproteinases (MMP) can actively digest the basement membrane. They are found to be up-regulated in inflammatory diseases, such as multiple sclerosis, which can compromise the integrity of the blood-brain barrier and contribute to disease progression.

Dystroglycan is a non-integrin receptor that mediates binding to proteoglycans and laminin within the basement membrane. This binding is essential for maintaining the structural integrity and function of the neurovascular unit.

Astrocytes contribute to the maintenance of the BBB by regulating its integrity. The loss of astrocytes is associated with a loss of BBB integrity in vivo. They secrete factors such as basic fibroblast growth factor, transforming growth factor beta, angiopoietin-1, glia-derived neurotrophic factor, and lipoproteins, which help maintain the BBB. Additionally, co-culture of isolated brain endothelial cells (BECs) with astrocytes or astrocyte-conditioned media results in reduced permeability and increased expression of transporter proteins like GLUT-1 and tight junction proteins.

BECs and astrocytes communicate through a two-way interaction, where BECs can influence astrocytic properties, and astrocytes can regulate the functions of BECs. This communication is essential for maintaining the integrity and functionality of the Blood-Brain Barrier (BBB).

Tight junctions (TJ) between BECs decrease paracellular permeability of the BBB to many molecules, especially ions, which contributes to the high electrical resistance of the BBB (1000-2000 ohms-cm²) compared to peripheral vessels (2-20 ohms-cm²).

BECs have a higher mitochondrial content (~10% of total cytoplasmic volume) compared to peripheral ECs (~4%), indicating higher levels of energy utilization in BECs.

ATP-dependent transporters in BECs transport blood-borne harmful agents back into the blood (e.g., P-glycoprotein) and help maintain ionic and nutrient homeostasis.

The main types of interendothelial protein complexes present on BECs at the BBB are adherens junctions and tight junctions (TJ).

Tight junction (TJ) proteins in BECs are regulated in both normal physiology and pathological states, making them highly dynamic.

Tight junction (TJ) proteins are composed of transmembrane proteins that span the membrane and bind to those on adjacent cells. They include junctional adhesion molecules (JAMs), occludin, and claudin family members. Cytoplasmic accessory proteins connect these transmembrane proteins to a pericellular ring of actin filaments, facilitating endothelial cell adhesion and paracellular permeability.

Extracellular mediators such as cytokines (e.g., TNF-, IL-1), oxidative stress (e.g., nitric oxide), inflammatory mediators (e.g., prostaglandins and histamine), and disease-associated cells or proteins (e.g., HIV virus, bacteria, amyloid beta) can induce a leaky BBB by regulating tight junction (TJ) translocation, decreasing TJ protein expression, or phosphorylating the TJ complex.

Intracellular signaling pathways that regulate tight junction (TJ) proteins include:

1. Protein kinase family (A, G, and C)
2. Mitogen-activated protein kinase family (MAPK, including JNK, ERK, and p38MAPK)
3. Small G protein family (especially rho and rac)
4. Wnt and eNOS pathways

Additionally, intracellular Ca2+ ions are key regulators of TJ protein distribution.

The main categories of BBB transport mechanisms include:

1. Passive diffusion
2. Efflux transporters
3. Solute carriers
4. Receptor-mediated transcytosis
5. Immune cell migration

Specific transport properties that further subdivide BBB transport mechanisms include:

• Na+-dependent facilitated diffusion
• Macromolecule transport

Lipid solubility is a key factor in determining the permeability of a substance through the BBB by passive diffusion. Substances that are more lipid-soluble can more easily cross the lipid bilayer of the endothelial cells in the BBB.

The primary route of entry for dissolved gases to and from the brain is diffusion across the plasma membrane, specifically for gases like O2 entering and CO2 exiting the brain.

The overall negative surface charge on the luminal membrane of Brain Endothelial Cells (BECs) due to heparan sulfate proteoglycans results in greater access for cationic molecules compared to anionic ones. This is important for designing therapeutic agents targeting the brain.

Solute carriers at the BBB allow access to essential molecules for metabolism, including glucose, amino acids, and some ions.

Glucose transport at the BBB is mediated by facilitative diffusion through glucose transporter-1 (GLUT-1), which is expressed symmetrically on both luminal and abluminal membranes.

GLUT-1 can mobilize an intracellular pool in times of increased neuronal demand, such as during seizures, to facilitate glucose transport.

The L1 system transports essential amino acids such as leucine, valine, isoleucine, phenylalanine, and tryptophan.

The Y+ system is a cationic transporter that facilitates the transport of lysine, arginine, and ornithine into the brain.

The systems expressed only on the luminal membranes are the glutamine system (n) and the acidic amino acid system (XG), which have high affinities for glutamine and glutamate, respectively.

The BBB regulates glutamate levels through the polarized control of glutamate transport. The excitatory amino acid transporter family (EAAT) uses the Na+ gradient to transport interstitial fluid (ISF) glutamate into brain endothelial cells (BECs). Glutamate then diffuses into the blood via system XG. This regulation is crucial to prevent excitotoxicity from excess glutamate.

Glutamate transport from plasma to the brain is virtually impossible due to the absence of a facilitative glutamate transporter and the high expression of excitatory amino acid transporters (EAAT) on the abluminal side of the BBB, which prevents glutamate from entering the brain from the plasma.

Glutamine transport across the BBB is regulated by Na+ transporters A and N, which pump glutamine into brain endothelial cells (BECs). Once inside, glutamine can either be transported into the blood via system N or converted to glutamate through the action of glutaminase.

Tight junctions (TJ) limit the free movement of ions across the BBB, thereby maintaining the integrity and selective permeability of the barrier.

The Na+-K+ ATPase is located on the abluminal BBB membrane.

The Na+/H+ exchanger is located on both the abluminal and luminal membranes and helps maintain ionic gradients and fluid movement across the BBB.

The luminal ion transporters include:

1. Na+/Cl- co-transporter - facilitates sodium and chloride ion transport.
2. Na+/K+/Cl- co-transporter - involved in the transport of sodium, potassium, and chloride ions.
3. Na+Cl-/HCO3- dependent influx transporter - aids in the influx of sodium, chloride, and bicarbonate ions.

Maintaining the Na+ gradient is crucial for the function of Na+-dependent transporters, which are essential for fluid movement and controlling ionic gradients across the BBB.

Na+ dependent symporters eliminate amino acids from the brain, preventing excess accumulation.

Facilitated diffusion allows essential amino acids (L1, Y+) and glucose (Glut-1) to enter the brain while eliminating excitatory amino acids (N, XG) into the blood across the luminal and abluminal membranes.

ABC transporters (P-gp, BCRP, MRP) protect the brain from toxins circulating in the blood.

Receptor-mediated transport allows essential proteins and signaling molecules, such as insulin and transferrin, to enter the brain and can also mediate the export of materials from the brain.

Adsorptive mediated transcytosis (AMT) allows cationic molecules access to the brain via non-specific transcytosis due to the net negative charge of the glycocalyx of the luminal BEC membrane, facilitating transport in both directions (brain-to-blood and blood-to-brain).

Receptor-mediated transcytosis (RMT) is the primary route of transport for essential peptides and signaling molecules across the BBB. It involves ligand-receptor binding, followed by endocytosis and exocytosis, allowing for the transport of molecules like insulin, leptin, and transferrin without lysosomal degradation.

Transferrin-bound iron is an important example of receptor-mediated transcytosis. Iron, which is essential for various biological functions, is bound to transferrin in plasma. It is taken up in endocytic compartments via the transferrin receptor, and under acidic conditions in the endosome, iron dissociates from transferrin and becomes available for uptake after exocytosis.

The steps involved in receptor-mediated transcytosis (RMT) at the BBB include:

1. Ligand-receptor binding: The target molecule binds to its specific receptor on the endothelial cell surface.
2. Endocytosis: The cell membrane engulfs the ligand-receptor complex, forming an endocytic vesicle.
3. Transport through the cell: The vesicle is transported across the cell.
4. Exocytosis: The vesicle fuses with the opposite membrane, releasing the ligand into the extracellular space.

This process bypasses the lysosomal degradation pathway, allowing for the direct transport of essential molecules.

ABC transporters on the luminal membranes of the BBB restrict brain entry of many molecules by using ATP to actively eject a range of substances, including xenobiotics and endogenous toxic molecules.

Lipid solubility is generally a good indicator of brain penetration; however, for some compounds like phenobarbital, brain penetration is much less than expected despite their lipid solubility.

P-gp contains two nucleotide-binding domains and 12 transmembrane segments. It is arranged in a barrel-like configuration with a 75-amino-acid linker region. Each half of P-gp includes a transmembrane domain (TMD) made of six transmembrane segments and an intracellular nucleotide-binding domain (NBD).

P-gp expression on the luminal membrane of the BBB plays a crucial role in protecting the brain by transporting various substances, including drugs, out of the brain, thereby influencing drug bioavailability and resistance to treatment.

ATP binding induces a conformational change in the transmembrane domain (TMD) of P-glycoprotein, which facilitates ligand translocation.

ATP hydrolysis leads to the dissolution of the closed nucleotide-binding domain (NBD) dimer, releasing ADP and allowing P-gp to return to a high-affinity ligand binding state.

P-glycoprotein substrates are generally hydrophobic and include endogenous molecules like steroid hormones and cytokines, as well as pharmaceutical drugs such as vinblastine, verapamil, and HIV-protease inhibitors.

The specificity of P-glycoprotein prevents many therapeutic drugs from reaching their target site, thus hindering their effective use.

Microglia develop and reside in the brain tissue, acting as immune-competent cells during development.

The brain is considered a relatively immune privileged site, with leukocyte traffic estimated to be 100 fold less than into the spleen or lungs, indicating lower immune surveillance.

Decreased CNS immune surveillance is mediated by the barrier nature of the cerebral vasculature and the restricted expression of cerebral cell adhesion molecules required for leukocyte capture from the blood.

During inflammatory conditions, there is a marked up-regulation of cell adhesion molecules, which can lead to neuronal damage in diseases such as multiple sclerosis.

Leukocytes are initially captured by P-selectin and vascular cell adhesion molecules (VCAM) on brain endothelial cells (BECs). Chemokines up-regulate integrins in leukocytes, allowing for firm binding and subsequent diapedesis through BECs (transcellular transport) and between cells (paracellular transport).

BBB dysfunction is found in conditions such as Alzheimer's disease (AD), multiple sclerosis (MS), and stroke.

Conditions that can compromise the BBB or TJ include brain tumors, inflammation, multiple sclerosis (MS), infections, trauma, and stroke.

BBB breakdown in acute brain injury can lead to:

1. Severe brain edema (increase in brain fluid volume)
2. Increased intracranial pressure
3. Potentially death.

In the initial stages after brain injury, the following changes occur in the BBB:

• Increase in the number of caveolae/vesicles that transport plasma proteins
• Decrease in the expression of tight junction (TJ) proteins
• Disruption of the actin cytoskeleton and the basement membrane (BM).

Cerebral ischemia is an event where parts of the brain do not receive sufficient blood supply to maintain neuronal function. It can lead to increased microvascular permeability and edema both clinically in vivo and in vitro.

Under hypoxic conditions, there is a reduced expression of occludin and an increase in brain permeability as observed in experimental models.