Conducting Neural Dissonance
Ion channel promotes neurodegeneration in mice
Inappropriate immune activity has occupied center stage in the study of MS, but clinical symptoms have turned a spotlight on the resulting neurodegeneration. A new paper, published online November 18 in Nature Medicine (Schattling et al., 2012), suggests that a particular sodium channel contributes to neuronal death in an animal model commonly used to study MS. Inflammatory damage to neurons can throw them out of whack, and the ion channel, TRPM4, seems to amplify the problem. This work might lead to therapies that directly counteract neurodegeneration—and an existing diabetes drug that blocks the channel already shows promise in mice. Experts say the findings justify a clinical trial in people with MS.
"It's a very solid and nicely worked-out story implicating an interesting ion channel" in MS-associated neurodegeneration, says Reinhard Hohlfeld, a clinical neuroimmunologist at the University of Munich in Germany, whose perspective on the paper appears in the December issue of Nature Medicine. "The findings are especially important for primary progressive and secondary progressive disease, which cannot be well treated."
The report of a new target and potential drug expands the idea that ion channels participate in axonal degeneration and that jamming them has a protective effect, says Raju Kapoor, a consultant neurologist at the National Hospital for Neurology and Neurosurgery in London, who was not involved in the study. "It's been known for over 20 years that sodium channels are overexpressed in demyelinated axons," he says. "The basic idea is that axons respond to injury by acquiring a complement of several different ion channels.” Some of these proteins trigger an “injury cascade” that renders neurons especially vulnerable to damage when loss of the myelin sheath leads to extra energy demands. The identities of the culpable ion channels—and of drugs that block them—are now emerging.
In the new work, Manuel Friese, a neurologist at the University of Hamburg in Germany, and his colleagues set out to look at the effect of TRPM4 on experimental autoimmune encephalomyelitis (EAE), a disorder in mice that mimics some aspects of MS (see "Experimental Autoimmune Encephalomyelitis"). They reasoned that the ion channel might be pathologically misregulated in MS and EAE, based in part on earlier studies that implicated a protein with a related function, the acid-sensing ion channel-1 (ASIC1), in EAE and in MS (Friese et al., 2007; Vergo et al., 2011). A drug that blocks ASIC1 reduced EAE-associated myelin damage, axonal degeneration, and motor problems in mice. What’s more, large quantities of ASIC1 appeared in portions of lesions that were active at the time of death in postmortem spinal cord and optic nerve tissue from nine patients with acute multiple sclerosis, but not in control tissue.
TRPM4—the full name is transient receptor potential melastatin 4—belongs to a large and diverse family of channel proteins. TRP channels form pores in the cell’s plasma membrane and offer passage to positively charged ions, such as sodium. In TRPM4, sodium ions can theoretically flow both ways, but scientists have reported the channels funneling sodium into cells, not out, says co-author Rudi Vennekens, a cell physiologist at the Catholic University of Leuven in Belgium. In mice, TRPM4 can limit histamine release during allergic skin reactions. The normal role of TRPM4 in people is unknown, but it appears in the heart, arteries, gastrointestinal tract, and immune system. On the pathological side, people in certain families with an inherited type of life-threatening arrhythmia have mutations in the protein. When Benjamin Schattling, a postdoctoral fellow in Friese's lab, was scouting for potential connections between ion channels and MS in the scientific literature, he noticed that the channel protein is activated by high intracellular calcium levels and needs ATP to close. In other words, TRPM4 turns on and stays on when calcium is abundant and ATP is scarce—conditions documented in MS.
As a step toward pursuing a possible link between TRPM4 and MS, Schattling and his colleagues induced EAE in mice that lack the channel protein and in wild-type littermates. The central nervous systems of both types of animals were brimming with the signs of inflammatory activity that typifies EAE, yet disease severity was unusually mild in mice missing TRPM4. The researchers wondered whether the protective effect maps to the absence of TRPM4 in immune cells, but several subsequent experiments suggested it does not. For example, they collected various immune cells that had infiltrated the central nervous systems of EAE-induced mice and found similar numbers in the wild-type and genetically engineered animals. They also replaced bone marrow—the source of all blood cells—from the TRPM4-lacking mice with that from wild-type animals and vice versa. Then they induced EAE. Donor bone marrow that was missing the ion channel did not protect wild-type mice from disease, nor did wild-type bone marrow that carried TRPM4 render the knockout mice susceptible. Lack of the protein must guard against EAE through a different cell type, the researchers concluded, and they turned their attention toward neurons.
To explore this idea, they exposed neurons grown in the lab to toxic conditions known to exist in MS: high glutamate concentrations; glucose depletion; or energy starvation, which was achieved by chemically inhibiting mitochondrial activity. Neurons that bore TRPM4 suffered significantly more damage than did neurons that lacked the protein. Extra calcium and sodium flooded the wild-type cells, which swelled to a larger volume than did those missing TRPM4. The team members wanted to verify that TRPM4 could explain these effects. Using patch clamp recordings on the mouse neurons, they measured the electrical activity of individual TRPM4 channels at rest and when stressed with extra glutamate. The results showed that current flowed inward under the influence of glutamate and not during normal cell conditions, which established that TRPM4 channels respond to pathological conditions that mimic those in MS.
All told, TPRM4's role seems consistent with the prevailing view of ion activity in the neurodegeneration of demyelinated axons, says Bruce Trapp, a neuroscientist at the Cleveland Clinic in Ohio. Removing myelin hobbles the superfast node-to-node transmission of nerve signals, forcing the axon to work harder to push each signal along (see "Altered Immunity, Crippled Neurons"). Each herculean effort drains the available ATP, fuel that normally would be used to pump out sodium and reset the axons to fire again. Instead, sodium accumulates, kicking into reverse a normally protective mechanism, the calcium-sodium exchanger. It usually extracts dangerously high calcium levels from inside the cell and slurps in sodium. When the sodium concentration is high, however, the exchanger swaps out sodium for more calcium instead, keeping TRPM4 turned on and leading to sodium overload. The neuron swells with water as it tries to dilute the sodium. In culture dishes and perhaps in the brain and spinal cord, "the cells increase in volume and then they die," Friese says. "It's almost a triple whammy," Trapp says, referring to the combination of extra glutamate, calcium-sodium exchanger reverse, and TRPM4 that can work together to damage the neuron.
To probe whether these findings might translate to MS, the researchers tested for evidence of TRPM4 protein and mRNA in postmortem brain samples of nine people with MS and four people without disease of the central nervous system. "In healthy brains, it's hard to find," Schattling says. In contrast, TRPM4 appeared in areas of the white matter that showed demyelinating activity from shortly before the time of death and not in older lesions.
Finally, the researchers asked whether a drug that blocks TRPM4 protects mice from EAE. They used glibenclamide (a generic drug also called glyburide in the United States), an oral medication for type 2 diabetes that is known for its effects on a different ion channel in pancreatic beta cells but that also thwarts TRPM4. This agent reduced EAE-associated symptoms and neurodegeneration in wild-type mice but provided no extra benefit to mice that lack TRPM4.
"If this work can be extrapolated to MS, it would suggest that blockade of the TRPM4 channels with drugs like glibenclamide may protect axons from degeneration, thus preventing progression of disability," wrote Stephen Waxman, a neurologist at Yale University who was not involved in the study, in an email. He added that the work augments the evidence that degeneration is triggered by inflammation but then proceeds independently in disorders such as MS (Waxman, 2006).
The accumulating evidence suggests, Hohlfeld says, that several ion channels—including ASIC1 and TRPM4—“may synergize in their neurodegenerative mechanisms." If so, targeting multiple channels simultaneously might provide a fruitful approach for combating MS. An appealing aspect of the growing array of ion-channel targets, Kapoor says, is the availability of approved drugs marketed for other indications, such as epilepsy, high blood pressure, arrhythmia, and now type 2 diabetes. Testing existing sodium-channel blockers in people with MS will likely take place in investigator-led trials, Trapp says, because drug companies lack the financial incentive to invest in expensive clinical trials of compounds that are available as generics.
So far, one clinical study has investigated whether a channel blocker helps individuals with MS. Based on mouse work similar to that reported in the current paper, Kapoor and his colleagues tested a sodium-channel blocker, lamotrigine, in a two-year, phase II clinical trial on 120 people (Kapoor et al., 2010). It failed to slow down secondary progressive MS according to the study’s primary clinical endpoint: cerebral volume as measured by brain imaging. But Kapoor points out encouraging secondary findings: Treated patients had only half the slowdown in a timed walk of 25 feet, and there was an apparent protective effect on neurofilaments, potential biomarkers of neurodegeneration that are shed into tissue fluids after axonal damage.
All approved MS drugs, as well as those furthest along in clinical testing, work mainly by beating back inflammation in the brain and spinal cord. "Unless you switch off inflammation that is driving the whole disease process, it's unlikely you're going to succeed" in foiling the disease, Kapoor says. Some medications may further slow disease by acting on the ensuing neurodegeneration, but evidence for that activity is sparse, say all the researchers interviewed for this story. Such capabilities are needed to fight progressive forms of the illness, for which no good therapies yet exist. "The current anti-inflammatories are working pretty well to slow [the] disease, but they don't stop it,” Trapp says. Perhaps the growing cast of ion channels will help identify compounds that could eventually deliver star neuroprotective performances.
Key open questions
- Assuming the scenario outlined in the story holds up in people, is aberrant TRPM4 activity associated with MS specifically, or does it (and perhaps other ion channels, such as ASIC1) contribute more broadly to neurodegenerative disease?
- What is the normal role of TRPM4 in axons?
- What causes TRPM4 to accumulate in demyelinating and injured axons?
- Does TRPM4 have a purely deleterious role in axon injury, or might it be beneficial in certain circumstances?
- Does glibenclamide reduce disease activity and/or progression in people with MS?
- Can TRPM4 blockers that are more selective than glibenclamide be identified? Will they be safe and effective in mice?
Thumbnail image on landing page. Reprinted by permission from Macmillan Publishers Ltd: Nature Reviews Immunology, Calcium signalling and cell-fate choice in B cells, Scharenberg et al., copyright 2007.