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Those of you who are reading this “blog” undoubtedly know that MH is an inherited disorder characterized by abnormal structure and function of a specific calcium channel within skeletal muscle. This channel, called the ryanodine receptor type one (RYR-1), is like a control valve that guides the release of calcium from the storage site in muscle, called the sarcoplasmic reticulum, into the inside of the cell, or myoplasm. When the cellular concentration of calcium increases, the muscle filaments that underlie contraction move together. Under ordinary circumstances the increase in cellular calcium is very transient. The excess is sopped up again by the sarcoplasmic reticulum and the muscle relaxes. When the calcium levels increase they also turn on a series of metabolic processes that provide energy to make the cellular filaments move, and power the pumps that take up the calcium again. Prolonged or repeated muscle contraction leads to not only energy production but also heat production (you know this when you exercise). Normally that is not a problem.
In the MH susceptible patient, there is an abnormality of the calcium channel structure, such that on exposure to gas anesthetics and succinylcholine, the channel stays open for a long time leading to prolonged increase in muscle cell calcium and energy production along with heat. If the process continues for a while leading to depletion of the energy stores, then the cell membrane begins to break down and various constituents such as potassium, the pigment myoglobin and the enzyme creatine kinase are released into the blood stream. Also heat is produced leading to an elevation of body temperature. So, what happens in MH is an accentuation of essentially a normal phenomenon. There is a failure of the spigot to turn off that releases calcium into the cell.
In most cases an abnormality in the DNA of the gene that is responsible for the manufacture of this channel may be found. As I mentioned in last month’s blog, in some cases, there may be a deficiency of another protein, Calsequestrin-1, that binds calcium so that excess calcium remains in the cell.
What about heat stroke? Heat stroke is a response to high environmental heat and can happen to just about anyone under the correct situations. The heat accelerates metabolism leading to exhaustion of energy stores and membrane breakdown. If the temperature is high enough the proteins in the cell begin to change their structure and “denature” and don’t function normally. For many years there has been a question as to whether MH susceptibles are more prone to heat stroke because, in the pig model for MH, the animals with the abnormal gene and abnormal ryanodine receptor upon exposure to high temperatures (and no other triggers) develop heat stroke as well as MH. There have been a number of patients who have developed life threatening heat stroke who were found to be MH susceptible on halothane and caffeine testing as well as genetic testing, but not all victims of heat stroke have been found to be susceptible to MH and certainly not all MH susceptibles develop heat stroke. In all honesty though, there really has not been more than a sporadic and limited study of the relation between MH and heat stroke. A more comprehensive study is sorely needed.
Last year however, a tremendous new insight into the possible relation between MH and heat stroke was found. This was a study conducted by a consortium of scientists including two MHAUS PAC members, Drs. Susan Hamilton of Baylor University, Robert Dirksen at the University of Rochester, mentioned in last month’s blog, and others. They had developed a mouse model for MH whereby one of the mutations known to underlie MH was installed in the gene for the RYR-1. Not only did the animals develop MH on exposure to the anesthetic gases, but they also developed MH when placed in a hot environment.
Studies of the mechanism whereby this occurs in Dr. Hamilton’s and colleague’s laboratory showed that these MH animals had a chronically elevated intracellular calcium level that led to the production of nitric oxide in the cell. Nitric Oxide(NO) is a mediator of many cellular processes including the inflammatory response. Under normal circumstances this is not a problem. However with heat, it was shown that more calcium is released and leads to formation of excess NO and a form of NO called a reactive nitrogen species. These reactive compounds bind to proteins in the RYR-1. When that happens, the channel in the MH mouse opens and behaves just as if an MH trigger anesthetic was present. Hence all the signs of MH occur!
Let’s turn to Duchenne Muscular Dystrophy (DMD) now. DMD is a tragic disorder whereby males (rarely girls) develop muscle weakness and wasting and usually die by their late teens. This is the disorder that led to the creation of the Muscular Dystrophy Association and for which Jerry Lewis raises large amounts of money each year.
A long time ago it became apparent that there was a genetic defect that was carried on the X chromosome. Males are characterized by having an XY chromosome. When there is a single dose of the gene on the X chromosome (males) the gene defect is expressed. When there is a double dose of the X chromosome (in the face of only a single dose of the defect), which is characteristic of females, the genetic defect is not expressed. In the 1980s it was found that DMD is characterized by an absence of a particular protein called dystrophin. This protein is a “structural “protein sort of like a scaffold for the cell. Without the benefit of this structural protein the muscle membrane develops tears and the integrity of the physiologic processes that maintain the entire cell structure is compromised. It was also noted that there is a chronically elevated calcium level in the muscle cells of the DMD patients. Presumably the calcium leaks into the cells through the tears in the membrane. Remember that in MH there is also an increased level of intracellular calcium, but it is not generally a chronic elevation.
In the 1990s case reports began to appear documenting that patients with DMD when exposed to MH trigger agents will often develop life threatening reactions characterized by a massive increase in blood potassium (leaking out from the muscle cell), to the point that the heart rhythm is disturbed and cardiac arrest occurs. In addition, massive muscle breakdown is noted and even occasionally temperature elevation. There however, did not seem to be a connection to MH. After all, DMD is associated with a structural protein whose gene is located on the X chromosome, and MH is associated with a change in the calcium channel associated with a completely different gene! Furthermore muscle biopsy testing using the caffeine halothane contracture test was inconclusive. The response often was not normal, but also not typical of MH.
However, a very recent report from the laboratory of Dr. Andrew Marks from the Department of Physiology and Cellular Biophysics at Columbia University suggests that the ryanodine receptor is indeed an important part of the pathophysiology of DMD. Using a mouse model of DMD, they found that these mice had a defect in their ryanodine receptors which allowed for binding to NO (a process called S-nitrosylation). S-nitrosylation of the RYR1 subsequently affects the ability of the receptor to bind to a protein called Calstabin-1. This chain of events causes a decrease in the channel’s stability and calcium leakage from the cell, leading to muscle damage and decreased muscle function... The NO derives from inflammatory cells that are found in and around the dystrophin deficient DMD muscle. Not a terribly dissimilar mechanism from heat stroke, but over a different time frame. The story is actually more complex than I have stated, but the bottom line is the same. So perhaps, here is the explanation of why the patients with DMD have a problem with anesthetics. It is because their RYR-1 receptors are abnormal due to the binding of the reactive NO compounds and in conjunction with the anesthetic gases there is an increased release of calcium into the cell leading to an MH like reaction. Now, since it is known that the cell wall is damaged in patients with DMD, therefore, perhaps there is a greater opportunity for the intracellular constituents such as potassium to leak out rapidly. (It should be mentioned that in MH an elevation of blood potassium often occurs too).
I probably have not done justice to the excellent, detailed work done by those who worked on the mouse model of MH and heat stroke nor on the work done with the DMD mouse (officially called the mdx mouse). I too am just learning about the intricacies of cell signaling and protein modification by small molecules.
It therefore turns out that the intracellular calcium channel, RYR-1, is probably involved in numerous cellular physiologic processes. Dr. Marks lab has also been studying the cardiac form of the RYR-1 , called RYR-2 and has good evidence that abnormalities in RYR-2 are associated with certain sudden death syndromes and even heart failure. Furthermore, they have identified a novel chemical that seems to prevent the processes I have described.
Therefore MH is one end of the spectrum of a wide variety of disorders effecting muscle structure and function. MH and the other disorders all alter the structure and function of the ubiquitous ryanodine receptor. I expect that we will soon learn that other disorders characterized by muscle breakdown and hyperthermia such as the Neurolept Malignant Syndrome will also display abnormal ryanodine function due to the production of intermediates such as reactive NO species.
Let’s however bear in mind that the studies I have cited have been conducted in non human species. It is well known that the genetic background of an animal does determine biologic responses. I am sure that this is the very beginning of a very exciting era in cell physiology and the central role of the RYR-1 channel.
Bellinger AM, Reiken S, Carlson C et al. Hypernitrosylated ryanodine receptor calcium release channels are leaky in dystrophic muscle. Nature Medicine 15:3:325-329, 2009
Durham WJ, Aracena-Parks P, Long et al. RYR-1 S Nitrosylation Underlies Environmental Heat Stroke and Sudden Death in Y522S RYR1 Knockin Mice.
Cell 133;53-65, 2008