Saturday, December 24, 2011

Brain Barriers


We like to think the brain is special. In fact, all the neurons and associated satellite cells throughout the mammalian central nervous system (CNS)—which includes the spinal cord—comprise a highly specialized tissue. The complex circuitry executed by the CNS requires a unique cellular environment. The circulatory system maintains the fluid bathing most bodily tissues, but the CNS inhabits its own cerebrospinal fluid (CSF). In addition, a poorly understood system of cell–cell connections defines a "blood–brain barrier" (BBB) that segregates the brain from the blood vasculature to maintain distinct local chemical conditions. BBB integrity is crucial for normal health, but it also presents a challenge to drug delivery. Although a variety of hereditary and age-related cognitive disorders, as well as certain brain cancers, may prove responsive to drug treatment, any potentially therapeutic chemical must bypass the BBB first.

The October 25, 2011, meeting of the Biochemical Pharmacology Discussion Group addressed Brain Barriers: A Hurdle for Drug Discovery and brought together biochemists, clinical pharmacologists, and industry specialists to explore current BBB research. Speakers considered the physiological nature of the BBB, emerging techniques to transiently disrupt the BBB or to allow drugs to circumvent it, and the risks associated with these techniques.

In her introductory remarks Jo Ann Dumin, an organizer of the Brain Barrier event and researcher at Satori Pharmaceuticals, highlighted the intersector nature of the field by calling for a non-competitive consortium between academia and industry to solve the BBB challenge. The nascent research presented throughout the day and the intersection of ideas born from multiple disciplines underscored her suggestion.

Joel Pachter of the University of Connecticut Health Center began by demonstrating that physiologists do not agree which level of vascular organization or subcellular structures designate the BBB. Definitions range from general, citing only endothelial cells lining blood vessels, to detailed, naming specific tight junctions that lash cells together. Pachter argued that rigorous biological knowledge of BBB construction must inform clinical work. This notion drives research in his laboratory where experimental systems have been devised to identify relevant components of the BBB. He showed that advanced fluorescent microscopy can be used to calculate the densities of tight junctions on different blood vessel surfaces. Capillaries top the list in this assay for tight junctions per unit surface area, suggesting the vessels are key to the BBB.

Pachter further explained that cultured endothelial cells derived from brain tissue and used routinely in laboratories may not accurately reflect the BBB, because the vascular population from which they originated cannot be readily defined. To get a better understanding of which vessel population(s) contribute to the BBB, he proposed using laser capture microdissection, which allows for selection of specific cells from a heterogeneous population. The technique can be used to analyze endothelia from distinct types of vessels separately, allowing for comparisons of global gene and protein expression patterns. Pachter concluded by emphasizing the perhaps underappreciated heterogeneity of endothelial tissues.

Echoing Joel Pachter's introduction, Damir Janigro of the Cleveland Clinic pointed out that even apparently trivial questions about the BBB remain unanswered. His talk pivoted around the effects of induced, and even pathological, BBB disruption. Janigro's team studies epilepsy and the associated seizures, among other neuropathological disorders. Their analyses of a rat model of epilepsy in which pilocarpine administration induces seizures revealed that the drug does not significantly penetrate the BBB under any conditions, but it does precipitate BBB disruption. Direct injection into the brain resulted in loss of consciousness without convulsions. These observations suggested that mere BBB breach may cause seizures. A battery of experiments in model organisms correlated BBB opening with seizure onset, as tracked by electroencephalography (EEG), a measure of electrical activity in the brain.

Janigro's group also tested the hypothesis in human patients undergoing clinical surgeries and found that elevated serum S100β—a molecular marker of a breached BBB—predicted seizure onset. These experiments informed clinical work by Janigro and his colleagues on certain forms of epilepsy, but the results are also pertinent to the development of intentional BBB disruption methodologies for the purposes of drug delivery. Janigro introduced the ex situ endothelial culture tubes used in his laboratory to screen for the effects of various drugs on BBB permeability. The plastic tubes are seeded with human endothelial cells and allowed to develop into "mock" blood vessels. By flowing buffer through the vessels (mimicking blood flow) the researchers can observe the effect of shear forces on endothelium, while the permeability of the cell layer serves as a bench-top model of the BBB.

The day's lectures centered on the endothelial barrier between blood and brain tissue, but Adam Chodobski from the Warren Alpert Medical School at Brown University spoke about the less appreciated blood–cerebrospinal fluid barrier (BCSFB). The BCSFB resides in the choroid plexus, a highly vascularized organ consisting of epithelial cells, which is located within the ventricles (CSF-filled cavities) of the brain and is responsible for generating CSF. Since this liquid bathes the brain and spinal cord, it too is segregated from blood. The separation is similar to but distinct from that characterizing the BBB, and therefore represents an additional facet of brain-targeted drug delivery. Chodobski pointed out that CSF, which turns over four times a day in an adult human and drains into the peripheral circulation, plays an important role in clearance of drugs from their target sites. As delivery routes, the BCSFB and CSF may be useful for targeting brain cells abutting the CSF space, but other brain regions may be too far for effective distribution of introduced chemicals.

David Begley of King's College London began by reiterating two themes that cut across all the talks. First, the endothelium of the BBB is not homogeneous; general descriptions of its molecular properties therefore fail to provide a complete picture. Second, CNS vasculature is distinct from peripheral vasculature. Specifically, peripheral capillary cells are fenestrated: not sealed together by tight junctions, which allows solutes, large molecules, and even cells to pass between them. Peripheral capillary cells also routinely transport certain molecules by engulfing them on one surface, and releasing them on the opposite surface. Current dogma holds that the BBB does not transcytose in this way, but Begley's research demonstrates otherwise. He and his collaborators observed that although receptor-mediated transcytosis across the BBB occurs relatively rarely, it nevertheless contributes non-negligibly to BBB permeability. The range of BBB-active receptors is poorly characterized, but at least some are known and appear to exhibit limited but still useful flexibility in the classes of molecules they recognize. This opened the door to a novel delivery method: nanoparticles. The engineered micron-sized beads are coated with a marker that docks to receptors mediating endocytosis. Initial trials have demonstrated that intravenously injected nanoparticles pass through the rodent BBB and end up in neuronal cytoplasm within 30 minutes. The beads degrade in the cell with time. Further pharmacological and materials science research will address precisely how the nanoparticles can serve as a platform for specific drugs.

A cell within the rat brainstem takes up a nanoparticle by receptor-mediated endocytosis. In his keynote presentation David Begley of King's College London highlighted endocytosis as a historically underappreciated route to bypassing the blood–brain barrier. (Image courtesy Anja Zensi, University of Frankfurt and presented by David Begley) 

Innovative technology also featured in the presentation by Elisa Konofagou. Her group at Columbia University experiments with focused ultrasound (FUS) as a noninvasive tool to transiently open the BBB. FUS targets brain regions selectively by disrupting only a small (mm2- to cm2-sized) defined area of the BBB, with most of the disrupting effects localized to the acoustic focus. As a high-frequency sound emitted by FUS propagates across living tissue, it imparts mechanical perturbations, which in turn generate heat. The secondary thermal effect can precipitate cell damage, but this can be minimized by an injection of microbubbles (already established and approved as safe in other applications). The microbubbles resonate with the applied FUS frequency and cause BBB opening without generating excessive heat.

MRI imaging of murine and primate brains during FUS revealed the size and duration of "passive cavitations" under various conditions, and the laboratory has honed in on a set of acoustic frequencies and microbubble sizes that generate cavities of desired area and duration. Because FUS physically breaches the BBB, it can be applied broadly to any drug that can be delivered intravenously. A graduate student in the Konofagou laboratory, Yao-Sheng Tung, presented his work on identifying the ideal microbubble size. Tung's poster received one of two prizes awarded in the symposium's poster competition.

Our molecular understanding of some CNS-associated disorders suggests that gene therapy, rather than drugs, may prove therapeutic. Gene therapy involves the targeted delivery of genetic material to cells otherwise lacking the correct genetic information. Getting genes into cells is challenging enough, but getting them past the BBB poses an even greater hurdle. Brian Kaspar of the Ohio State University and The Research Institute at Nationwide Children's Hospital studies the pathology and therapeutics of neuromuscular disorders, which often stem from problems behind the BBB. Kaspar and his colleagues have found that adeno-associated virus 9 (AAV9) can bypass the BBB. Since viruses are essentially small capsules capable of delivering genetic payloads to very specific targets, this observation opens many avenues for future research. Preliminary studies have demonstrated the feasibility and tolerability of the approach in non-human primates.

Conor P. Foley, one of two winners of the poster competition, described arterial delivery of gene therapy in mice: injection of a viral vector into a specific artery feeding only a target region of the brain, coupled with chemically-induced BBB disruption delivered high doses of the vector without the need for surgery.

William H. Frey II of the Health Partners Alzheimer's Research Center at Regions Hospital and the University of Minnesota spoke of avoiding the BBB altogether when targeting drugs to the CNS. Extracellular pathways connect the nasal passage essentially directly to the brain cavity in the region where olfactory nerves communicate between the two compartments. Intranasal delivery is therefore noninvasive, amenable to a wide range of chemical compounds, and functional with low drug doses. An interest in low glucose uptake by brain tissue in Alzheimer's patients led Frey and his colleagues to successfully demonstrate the effectiveness of intranasally administered insulin. More recently, intranasally administered stem cells in a rat model of neurodegenerative disease were shown to migrate into the CNS, with manifest possibilities for future clinical research.

Deep brain surgery brings fresh hope to sufferers

Consultant neurosurgeon Ludvic Zrinzo (right) testing the deep brain stimulation procedure on Andrew Falzon, one 0f the first five patients to undergo the groundbreaking surgical procedure in Malta.

Maltese neurosurgeon Ludvic Zrinzo and his team at a London hospital are revolutionising an established surgical procedure, bringing new hope to sufferers of neurological disorders.
The first Maltese patients underwent this procedure – deep brain stimulation (DBS) – at Mater Dei Hospital in July and the technique is now also being applied to British patients with severe refractory Tourette syndrome.
Tourette is a condition that causes involuntary tics including movements or sounds that cannot be controlled.
In a case that was all over the British media yesterday, an English female sufferer of the Tourette syndrome was fitted with a “brain pacemaker” to rid her of debilitating muscle movements that almost drove her to suicide.
The relentless muscle jerks made it difficult for her to breathe and walk and stopped her from cooking, driving and reading.
The pacemaker allowed her to return to normal life, she told the media.
Mr Zrinzo, the 39-year-old doctor who leads these surgeries at London’s National Hospital for Neurology and Neurosurgery, insisted that DBS did not provide a cure but could offer significant improvements in symptoms and the quality of life of both patients and their families.
“There is solid evidence that deep brain stimulation improves symptoms and quality of life in conditions such as Parkinson’s disease, dystonia and tremor. Research has shown promise in treating severe cases of some chronic pain syndromes and obsessive-compulsive disorder.
“So far, we have encouraging results on Tourette’s patients,” he said.
Claims that the treatment might be a placebo were quelled when a patient who had been installed with a brain pacemaker two years ago suddenly experienced a re-emergence of the symptoms when the batteries of the device were exhausted.
Similar studies with DBS are being carried out to treat cluster headaches, which are described by sufferers as the equivalent of having ahot iron rod thrust into their eye.
In severe cases, the headaches can last up to three hours and patients can get up to 10 attacks in a day.
Such a headache, estimated to affect one in 1,000 people, is unresponsive to medication in 10 per cent of patients.
“Considering that these patients had tried everything else and remained resistant, the reaction to the treatment we keep seeing is very encouraging,” Mr Zrinzo said.
The procedure is similar to a heart pacemaker, where electrodes are implanted deep within the brain and connected to a neurostimulator that lies underneath the skin on the chest wall.
The electrodes in the brain are connected by wires that pass underneath the skin and are connected to the pacemaker implanted on the chest wall.
The computer inside it then allows doctors to change the way the brain works, treating symptoms, most commonly those of Parkinson’s disease, such as tremor, rigidity, stiffness, slowed movement and walking problems.

Multiple sclerosis `linked to different areas of brain

Multiple sclerosis `linked to different areas of brain`
Washington: Scientists claim to have found evidence that multiple sclerosis affects an area of the brain that controls cognitive, sensory and motor functioning apart from disabling damage caused by the disease`s visible lesions.

A team at the University of Texas selected the thalamus of the brain as the benchmark for its research published in the latest edition of `The Journal of Neuroscience`.

"The thalamus is a central area that relates to the rest of the brain and acts as the `post office`. It also is an area that has the least amount of damage from lesions in the
brain but we see volume loss, so it appears other brain damage related to the disease is also occurring," Khader Hasan, who led the team, said.

Researchers have known that the thalamus loses volume in size with typical aging, which accelerates after age 70. The team`s purpose was to assess if there was more volume loss in patients with multiple sclerosis, which could explain the dementia-related decline associated with the disease.

"Multiple sclerosis patients have cognitive deficits and the thalamus plays an important role in cognitive function. The lesions we can see but there is subclinical activity in multiple sclerosis where you can`t see the changes.

"There are neurodegenerative changes even when the brain looks normal and we saw this damage early in the disease process," the scientists said.

For their research, the scientists used precise imaging by the powerful 3 Tessla MRI scanner to compare the brains of 109 patients with the disease to 255 healthy subjects.

Adjusting for age-related changes in the thalamus, the patients with multiple sclerosis had less thalamic volume than the controls. The amount of thalamic loss also appeared to be
related to the severity of disability.

"This is looking at multiple sclerosis in a different way. The thalami are losing cellular content and we can use this as a marker of what`s going on. If we can find a way to
detect the disease earlier in a more vulnerable population, we could begin treatment sooner," Hasan said. 

Thinner brains linked to Alzheimer’s disease: Study

More and more studies report of psychiatric problems being associated with abnormalities in the brain. Affirming this further, scientists from the American Academy of Neurology (AAN) have put forth that individuals not manifesting any memory problems at present, but having smaller portions related to the brain’s cortex may be at risk for early stage Alzheimer’s disease.

In this study, about 159 persons devoid of dementia aged 76 on an average were exposed to MRI scans to gauge the width of brain parts associated with the cortex. According to the outcomes, 19 patients with smaller size of certain parts related to the cortex were reported of being in the high risk group.

While nearly 116 fell into the moderate risk category, 24 of them were considered to be at low risk for the disease. Further, the subjects were also asked to fill in tests for comprehending their memory, problem solving capabilities and attention spans. Of the individuals in the high risk group, approximately 21% seemed to succumb to cognitive decline in the 3-year follow-up period. On the other hand, almost 7% in the average risk group and apparently no one from the low risk group experienced cognitive impairment during these years.

“Further research is needed on how using MRI scans to measure the size of different brain regions in combination with other tests may help identify people at the greatest risk of developing early Alzheimer’s as early as possible,” cited study author Bradford Dickerson, MD, of Massachusetts General Hospital in Boston and a member of the American Academy of Neurology.

Another finding of the study was that 60% of the people falling into the high risk group appeared to possess abnormal proportion of proteins in the cerebrospinal fluid, which is another hallmark of Alzheimer’s. This percentage was 36 among the average risk set and 19 amidst the lower risk group. The team has plans of expanding the study by close observation of MRI scans hereafter.