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RESEARCH BRIEFSMechanism Described that Links Migraine Aura and PainWhat causes migraine? The three membranes of the meninges--the tough dura mater, the arachnoid mater, and the pia mater--are permeated with blood vessels and layered to form a protective cap, one of the few areas of the brain that is wired to sense pain. Vasodilation in these tissues clamps the migraine sufferer in a hood of misery. But the processes connecting blood flow to neural-triggering events are poorly understood. In a study published in the February Nature Medicine, co-lead authors Hayrunnisa Bolay, Uwe Reuter, and Andrew Dunn, working with Michael Moskowitz, HMS professor of neurology at Massachusetts General Hospital, identify the neural basis for the long-suspected link between vasodilation, electrical activity in the cortex, and transmission of pain. This work provides the clearest evidence yet that "aura," the visual disturbance preceding migraine, is mechanistically related to headache pain.
Migraine pain begins in the cortex. Cortical spreading depression provoked by a pinprick to the rat cortex causes vasodilation and inflammation in the meninges and migraine pain. The process works on the brain circuit shown above: blood vessels (red circles) in the meninges are innervated by neurons from the trigeminal ganglion and the sphenopalatine ganglion (TGG and SPG, dark gray cell bodies). These communicate with the trigeminal nucleus (TGN) and the superior salivatory nucleus (SSN) in the brainstem. The circuits for migraine pain are remarkably similar in man and rat (see figure). The meninges are innervated by neurons from the trigeminal ganglion, located just above the palate. From there, bipolar cell bodies send out axon collaterals that terminate near blood vessels in the pia and dura mater. Other axons extend to the trigeminal nucleus in the brainstem. These impinge upon neurons that send signals to the rostral brain area, leading to the perception of pain. But what triggers the circuit? Moskowitz and others suspected that the answer might be found in a phenomenon called cortical spreading depression. This wave of electrical activity arises spontaneously in humans and can be elicited in animals, like the rats used in this study, by a pinprick to the cortex. Neurons in the resulting propagating wave dump potassium ions and neurotransmitters, and suck up water, setting off an ionic imbalance and prolonged suppression of neural activity. Moskowitz and colleagues used laser-speckle imaging to simultaneously measure blood flow in the middle meningeal artery, the major vessel supplying blood to the dura mater, and in the underlying vessels of the pia and cortex. They observed biphasic dilation of the dural artery provoked by spreading depression. During the early dilatory phase, which occurred as the cortical wave passed, there was also a spike in blood flow in the vessels of the cortex and pia. The researchers showed that noxious ions and metabolites released during cortical spreading depression sensitize the trigeminal axon collaterals impinging on the blood vessels in the cortex and meninges and set off a series of reactions. Axon terminals in the dura release neuropeptides and are directly responsible for a migraine-associated inflammatory response. This local activity is distinct from events that lead to brainstem activation. In the latter case, signals transmitted by irritated meningeal axons are relayed back to trigeminal ganglion bipolar cell bodies, then on to the brainstem. Here, in addition to activating rostral brain areas responsible for pain perception, the neurons triggered signaling to the superior salivatory nucleus and the sphenopalatine ganglion. Neurons from this ganglion synapse on the vessels of the dura mater and release vasoactive peptides, resulting in the second, prolonged dilation of the dural artery. About 20 percent of migraine sufferers experience the flashing, moving lights of aura for 20 to 40 minutes prior to headache. The accumulated data, most recently from positron emission tomography and magnetic resonance, suggests that aura and cortical spreading depression are aspects of the same phenomenon. The current study thereby links the sensations experienced by those afflicted to physiological events in the cortex. --Anne Mahon
Gene Therapy Technique Restores Function to Heart CellsA study led by researchers at Massachusetts General Hospital and HMS shows promise for a gene therapy that may help restore cardiac function in patients with heart failure. Abnormal calcium cycling in heart muscle cells causes heart failure in aging patients by interfering with the cells' normal cycle of contraction and relaxation. The MGH team, including principal investigator Roger Hajjar, HMS assistant professor of medicine, and first author Federica Del Monte, HMS research fellow in medicine, earlier showed that cardiomyocyte function could be restored in vitro using a gene transfer technique that causes overexpression of the gene for a calcium pump called sarcoplasmic reticulum calcium ATPase (SERCA2a; see Focus Research Briefs, Dec. 17, 1999). In the new study, which appears in the Feb. 26 Circulation, they used a similar strategy, employing an adenovirus vector to transfer genetic material into cells cultured from nine failing hearts removed from transplant patients and 18 nonfailing donor hearts as controls. But this time, instead of the gene for SERCA2a, they introduced antisense DNA that binds to and blocks the activity of the RNA message for phospholamban, a protein that regulates SERCA2a activity. Results were similar to the earlier SERCA2a gene transfer study. In both healthy and failing cardiomyocytes infected with the antisense DNA, phospholamban expression fell by more than 50 percent. In the failing cells, this decline increased the degree and speed of contraction, indicating improved functioning. Ultimately, the researchers hope the treatment strategy can be used to correct heart failure and cut down on the need for transplants. Preclinical trials are under way to test it in animals.
Method Advanced for High-Throughput Protein PurificationFor the burgeoning science of proteomics--defining the functions and interactions of an organism's entire set of proteins--the speed of progress will largely depend on how quickly these proteins can be made and purified.In the March 5 Proceedings of the National Academy of Sciences, researchers in the HMS Institute of Proteomics describe promising studies aimed at developing methods for high-throughput purification of human proteins for use in biochemical assays such as protein microarrays. The bacterium E. coli has become a convenient and inexpensive vehicle for growing individual human proteins. But different proteins require different conditions for their expression and purification. So a major challenge for proteomics is to find the best ways to express and purify thousands of proteins in parallel. Proteomics Institute director Joshua LaBaer, HMS instructor in medicine, with lead author Pascal Braun, research associate in medicine at Massachusetts General Hospital, and co-authors selected a test set of 32 human genes of varying size, expressed them in E. coli, and attached each product to four different "purification tags": hexa-histidine (HIS6-), calmodulin-binding peptide (CBP-), glutathione-S-transferase (GST-), and maltose-binding protein (MPB-). They analyzed the resulting 128 proteins for yield, purity, and losses. The larger tags, GST- and MPB-, produced the best results, yielding at least 300 ng/ml of purified protein from 26 of the 32 genes. In general, smaller proteins were expressed at higher levels than larger ones. To assess whether the proteins were active, they tested two of them in biochemical assays, where both showed activity. Applying their purification methods to 336 randomly selected human genes, the investigators were able to successfully purify 60 percent, a figure they believe represents a reasonable lower estimate for the overall success rate of this strategy. "The availability of such methods will alleviate a key bottleneck in the application of new proteomic techniques such as protein arrays to human biology," the authors write. --Two briefs above by Tom Reynolds |
