Cause and mechanism of homocysteine neurotoxicity: A role for glutamate hyperactivity.
Homocysteine is an amino acid containing sulphur, however this amino acid cannot be used as a building block for proteins, instead it is used for other important roles such as methylation. As it cannot be obtained by diet, it is produced by the demethylation of methionine which involves several steps. Firstly methionine is adenylated to S-adenosylmethionine (SAM) by the enzyme SAM synthase. SAM is then converted, by the loss of a methyl group, to S-adenosylehomocystein which is then hydrolysed to give homocysteine. Homocysteine then needs to undergo one of two pathways; re-cycling or elimination5.
In this way homocysteine is converted back to methionine by methylation of homocysteine. This is done by the methylcobalamin vitamin B12 dependant enzyme- methionine synthase which transfers a methyl group from 5-methyltetrahydrofolate to homocysteine converting it back to methionine. This recycling pathway is dependent on both folate and vitamin B12 in the diet and if enough is available, half of all homocysteine produced follows that pathway5.
In this pathway homocysteine is converted to Cysteine. It is firstly converted to cystathionine by the vitamin B6 dependant enzyme cystathionine β-synthase, which is then hydrolysed to for cysteine. SAM is responsible for the regulation of this pathway where maintaining blood cysteine is thought to play a role in regulation brain glutathione concentrations4,5.
Homocysteine plays a role in mechanisms in the brain such as the synthesis and degradation of neurotransmitters, membrane phospholipids and in methylation of DNA, RNA and proteins4,5. However, as viewed above, enzymes involved in the catabolism or recycling of homocysteine are strongly dependent on factors such as vitamin B6, B12 and folate and a deficiency of these or a mutation of the enzyme could cause a disregulation or abnormal amounts of homocysteine. This is thought to be the cause of neurodegenerative disorders.
Normal blood levels of homocysteine are about 10µmol/L (may vary with age)1. An increase of homocysteine levels is called hyperhomocysteinemia and this in turn can be the cause or contribute to neurodegenerative disorders5. There are several reasons for how this high homocysteine can occur and accumulate.
* Enzyme mutations
One of the reasons for this could be due to a defect in the enzymes required for the metabolism of homocysteine. Such defects can be at a genetic level (DNA mutation)or post translational rendering the enzyme incapable of function.
Another could be due to co-factors, such as vitamin B12, B6 and folate, being absent. If any of these are missing the enzyme cannot function.
For example, folate is required for the conversion of methionine to SAM. A deficiency in folate causes a depletion in SAM and hence a depletion in homocysteine6.
SAM/homocysteine is important in methylation. If there is a decrease in SAM there is less methylation, reduced DNA repair leading to genetic mutations which can cause apoptosis. A reduction in folate also causes an accumulation of homocysteine within the cell as folate is used in its metabolism6. The cell excretes the homocysteine to protect itself however this now exposes nearby cells to high levels of homocysteine5. These high levels of homocysteine effect the glutamate receptors such as NMDA leading to neurodenerative and psychiatric disorders6.
The NMDA (N-methyl D-aspartate) receptor is an ionotropic glutamate receptor. It requires two ligands for activation: glutamate and glycine. Upon voltage-dependant activation the ion channel opens by the release of Mg2+ (used to block the channel) and facilitates the transport of cations such as Na+ and Ca2+.
In the presence of low concentrations of glycine(10µmol/L), homocysteine acts as an antagonist to the NMDA receptor at the glycine site and hence this inhibits the action of the receptor. However if the concentration of glycine is high and the homocysteine levels are normal, or if there is a high concentrations of homocysteine (100µmol/L) and normal levels of glycine, there is an observed excitotoxic effect. Homocysteine in this case now acts as an agonist to the NMDA receptor at the glutamate site4. This causes the overactivation of the NMDA receptor, causing an influx of calcium ions and reactive oxygen species (ROS) within the neuronal cells. An accumulation of the ROS causes damage to the cells, such as DNA damage, and this triggers apoptosis1.
To note that although homocysteine is toxic, it is considered to be quite weak compared to homocysteic acid which is formed by the spontaneous oxidation of homocysteine ( which participates in redox reactions). Homocysteine acid becomes neurotoxic as it reaches levels equal to 0.3-0.5mmol/L unlike the higher amounts of homocysteine required to cause the same effect. Homocysteine acid, like homocysteine, activates the NMDA receptor causing an influx of firstly calcium ions and then ROS which accumulates in the neuronal cytoplasm. These free radicals cause an externalisation of phophatidylserine to the membrane which then initiates apoptosis1.
All these different events lead to a depletion of neurons causing many diseases.
Diseases caused by homocysteine neurotoxicity
In cultured neurons, homocysteine has shown to damage and kill the cells by inducing breakage in DNA. This is probably due to the impaired methylation in DNA. ATP reserves in the cell begin to deplete as they try to repair the DNA damage. This decrease in ATP along with oxidative stress caused by homocysteine is thought to be a factor in the neurodegenerative disorders- Alzheimer's, Parkinson's and Huntington's disease6.
The effect of stroke
During stroke or head trauma there can be damage to the blood brain barrier. This can cause exposure of plasmid amino acids to the brain e.g. glycine and homocysteine. In this way there is a higher risk of disease by the increased glycine and homocysteine levels in the brain3.
Alzheimer's disease can be caused by several different factors which include, genetic deposition, amyliod β-peptide accumulation, oxidative stress calcium ion mismetabolism and excitotoxicity.
Patients with this disease show high levels of homocysteine and it is thought that this may have occurred to before the onset of Alzheimer's and contributed to the disease. These patients have a reduction in SAM and a reduced activity of the enzyme used for its regeneration. Homocysteine can induce AD by its toxic effect of increasing calcium ions and ROS, contributing to oxidative stress and excitotoxicity. The oxidative stress can induce Aβ amyloids formation and this deposits in the brain causing further apoptosis6.
Like AD patients, PD patients have an increase in plasma homocysteine levels. It is thought that the current treatments of PD might actually be accelerating the disease. The current treatment of levodopa might promote the decrease in methyl groups and this in turn can cause and increase in homocysteine. This in turn, as described before will damage or kill the dopaminergic neurons causing further progression of the disease6.
As there are many factors leading to these disease many different targets that can be used to cure or slow down the disease process.
1) Targeting the NMDA receptor .
By using specific inhibitors such as MK-801 and α-methylcarboxyphenyl glycine, to the NMDA receptor causes a suppression in the effect of homocysteine. MK-801acts as an antagonist to the glutamate site on the NMDA receptor and so it competes with the homocysteine. By acting as an antagonist, the receptor will be turned off and this will reduce the influx of calcium ions and ROS into the cell protecting it from apoptosis.
Α-methylcarboxyphenyl glycine is an inhibitor to metabotropic glutamate receptors( group 1) which is responsible for inducing calcium ion release in the endoplasmic reticulum. This will hinder the calcium ion influx effect of homocysteine1.
2) Diet modification
Folate: since a deficiency on folate increases homocysteine levels, an increase of folate in the diet will reverse the effects5. Folate as described before is essential to the enzymes required for homocysteine metabolism. It has been shown that since the government has implemented folate supplements and its presence in processed foods, there has been a decrease in neural birth defects. Amounts of about 2mg of folate per day has shown to be effective in decreasing plasma homocysteine levels over time. Similarly vitamin B12 and B6 supplements could reduce homocysteine levels as they also are important for its metabolism. If taken on a regular basis it could reduce the risk of neurodegenerative and psychiatric disorders6.
As both homocysteine and homocysteic acid cause an accumulation of ROS, a mechanism to try to regulate ROS levels and protect the cell are being tested. For example Carnosine, a dipeptide which supports neurons when under oxidative stress. It functions by protecting the cell from damage caused by excitotoxic effect of the NMDA receptor. It is a good administered drug as if it is added to the blood it does in fact accumulate in the brain, lower toxicity and preventing neuron death. Also it is quite safe as even when overdosed it is hydrolysed by serum and kidney carnosinase which can prove effective against side effects1.
In conclusion, homocysteine can cause toxicity by both reduction in methylation and hyperactivation of the NMDA receptor. It causes influx's of calcium ions and ROS causing cell disruption and oxidative stress. This can lead to the damage and destruction of neurons leading to or adding to diseases such as Parkinson's and Alzheimer's disease. The effects of these can be reduced to taking drugs or vitamin/folate supplements, however if it is de to genetic defect rather than a deficiency, the effects will be minimal.
Even if drugs are used to target the NMDA receptor it does not stop the reduction in methylation.
1. Boldyrev A. (2009) Molecular Mechanisms of Homocysteine Toxicity. Biochemistry 74(6): 589-598.
2. Ho P, Ortiz D, Rogers E, Shea T. (2002) Multiple Aspects of Homocysteine Neurotoxicity: glutamate Excitotoxicity, Kinase Hyperactivation and DNA Damage. Journal of Neuroscience research 70L 694-702.
3. Lipton S, Kim W. Choi Y, Kumar S, D'Emilia D, Rayudu P, Arnelle D, Stamler J. (1997)Neurotoxicity associated with dual actions of homocysteine at the N-methyl-D-aspartate receptor. Proc. Natl. Acad. Sci. USA 94:5923-5928
4. Obeid R, Herrmann W. (2006) Mechanisms of homocysteine neurotoxicity in neurodegenerative disease with special reference to dementia. FEBS Letters 580: 2994-3005.
5. Martignoni E, Tassorelli C, Nappie G, Zangaglia R, Pacchetti C, Blandini F. (2007) Homocysteine and Parkinson's disease: A dangerous liaison? Journal of the Neurological Sciences 257: 31-37
6. Mattson M.P, Shea T.B. (2003) Folate and homocysteine metabolism in neural plasticity and neurodegenerative disorders. TRENDS in Neurosciences 26(3): 137-146.