Methylene Blue: A Novel Nootropic?
Neurons are highly metabolically active and rely on mitochondrial aerobic metabolism to carry out complex functions.
Mitochondrial dysfunction is associated with impaired memory and neurodegeneration. Methylene blue has been identified 1 as the forerunning candidate drug to improve mitochondrial function and ATP synthesis in neurons.
The nootropic effects of methylene blue are related to its ability to enhance memory consolidation and improve oxygen consumption in the brain.
A study in the late 70's concluded that methylene blue specifically enhances memory consolidation as opposed to memory retrieval or encoding in an inhibitory avoidance test in rodents 2.
These results were subsequently replicated in 2004 when administration of 1 mg/kg Methylene Blue USP to rats improved memory consolidation in a spatial task for food search 3.
Low-dose Methylene Blue also rescued memory impairment caused by treatment with cytochrome c oxidase inhibitors. Cytochrome c participates in the electron transport chain and is essential to energy generation in neurons. Inhibiting this protein therefore impairs mitochondrial bioenergetics and depletes cellular ATP, resulting in memory impairments. This result is at least suggestive that methylene blue’s nootropic effects involve mitochondrial enhancement.
Remarkably, it has also been reported that methylene blue dose-dependently increases oxygen consumption in the brain 4. The authors also noted that the range of doses which increased cerebral oxygen utilization corresponded to the same dose range that improved memory retention in avoidance and appetitive tasks.
Methylene Blue and Depression
Wager-Smith et. al. argue that depression and chronic, unpredictable stress cause synaptic wear-and-tear or “neuronal microdamage” that can sometimes escalate into frank neuroinflammation if left untreated. The authors cite the fact that chronic stress reshapes the morphology of the brain and the role that cytokines and other inflammatory mediators play in the pathogenesis of depression.
For a number of reasons, Methylene Blue may be a good candidate drug to hold this pernicious synaptic remodeling at bay:
- MB inhibits abnormal tau aggregation, implicated in neurodegenerative disease. Tau is a protein that stabilizes microtubules that help shuttle nutrients and other cargo in the neuron.
- MB is a potent antioxidant which readily accumulates in neural tissue.
- MB is an artificial electron acceptor/donor in mitochondria and therefore supports ATP synthesis.
- MB weakly and reversibly inhibits Monoamine Oxidase A. MAO inhibition has been used for decades as a therapeutic and potentially disease-modifying strategy in the management of Parkinson’s disease.
- Finally, MB has an excellent safety profile and is effective in such low doses that toxicity is highly unlikely.
Methylene Blue is most appropriate for the purposes of neuroprotection, down-regulation of pro-apoptotic pathways (apoptosis is orchestrated cell death), and support of endogenous antioxidant defense and replenishment of ATP. Though individuals with some synaptic wear-and-tear (so-called reversible “microdamage”) stand to gain the most from methylene blue, there is reason to believe methylene blue confers benefits in healthy subjects as well.
Dosage and Safety
TauRx’s clinical trial of Rember (a purified form of methylene blue)
used a dose of 50 mcg, which is the dose I recommend for the purpose of neuroprotection in this article.
A reader has helpfully pointed out that the above information is incorrect; the Rember trial used doses from 30mg – 90mg. I’m recommending a dose of 0.16 mg/kg, which is the lower boundary of the dose range suggested in this reddit thread.
Bare in mind that some individuals may wish to titrate a higher dose to also benefit from Methylene Blue’s Monoamine Oxidase-A (MAO-A) inhibition, which protects neurons by setting a ceiling on the hydrogen peroxide (H2O2) released by MAO during the natural metabolism (deamination) of serotonin. (H2O2 promotes the formation of free radicals). It is ill-advised to take an excessive dose, however, due to the hormetic inverted u-shaped dose response curve of methylene blue.
Methylene Blue has an excellent safety profile and has been used for decades with few adverse events at much larger doses than 13 mg to treat everything from malaria to cyanide intoxication. Despite this safety record, one should exercise caution because the synthesis of methylene blue involves the use of potentially neurotoxic reagents, including heavy metals like arsenic and organic solvents like dioxane.
One can avoid the inadvertent ingestion of toxic reagents by purchasing >99% pure, pharmaceutical grade, USP-compliant methylene blue, which can be obtained very cheaply and easily online. Aquatic disinfectants like Kordon Blue and reagent grade methylene blue intended for use in biochemical analysis is not appropriate for this purpose. USP-grade methylene blue, by contrast, conforms to United States pharmacopeia (USP) standards which ensures that the concentrations of contaminants are below unsafe thresholds, e.g., arsenic must be assayed below 8 ppm.
Methylene Blue Dosing Protocol
For convenience, I prepared a solution of methylene blue so that ten drops of solution would contain 13 mg of methylene blue. Since 10 drop is roughly equivalent to 0.5 mL, and the total volume of my empty dropper bottle is 30 mL, we obtain the following:
Desired dose: 13 mg
Volume of ten drops: 0.5mL
Dropper bottle volume: 30 mL
(13 mc / 0.5mL) = 26 mg/ml (desired final concentration of MB)
(26 mg/ml) * 30 mL = 780 mg = 0.78 g of methylene blue
Hence, I will need to dissolve 780 mg of methylene blue in a volume of 30 mL distilled water (the volume of my dropper bottle) to attain a final concentration of 26 mg/mL, so that ten drops of solution will contain about 13 mg +/- 5 mg.
- Pharma grade methylene blue USP
- Dropper bottle ($1 on Amazon)
- Distilled water (89 cents at Target)
- Scale/weighing paper/scoop
Bacteriostatic water is not necessary to reconstitute methylene blue due to methylene blue’s intrinsic antimicrobial properties. Hence, distilled water will be sufficient. (Methylene blue is used in the management of septic shock due to its ability to kill bacteria.)
I weighed out roughly 780mg of methylene blue.
And carefully added the methylene blue to my dropper bottle, along with ~30 mL of distilled water. Here’s the finished product:
Adding 10 drops to a blueberry smoothie is a fun way to administer this drug.
Methylene Blue's Mechanism of Action: Role for Mitochondria
Methylene blue has been used medically for decades as an FDA-approved treatment for methemoglobinemia (elevated blood concentrations of an oxidized form of hemoglobin) and urinary tract infections.
Impaired mitochondrial respiration and oxidative metabolism likely plays a prominent role in the pathogenesis of Alzheimer’s disease 5 and other neurodegenerative diseases. (The mitochondria are the powerhouses of the cell which produce the energetic currency ATP or adenosine triphosphate).
Dysfunctional mitochondrial bioenergetics produces a myriad of deleterious downstream consequences. Maintenance of the sodium-potassium ion gradients by the NA/K ATP-ase is extremely energetically demanding, requiring the constant hydrolysis of ATP and an uninterrupted efflux of ATP molecules across the outer mitochondrial membrane and into the cytoplasm.
It takes a lot of ATP to maintain the negative membrane potential that allows a nerve impulse or action potential to propagate down the axon. Any impairments in ATP production in neurons – the functional unit of the brain – will result in a deterioration in this membrane potential and result in pathological, spontaneous depolarizations.
Methylene blue is postulated to possess memory enhancing and neuroprotective effects in vivo related to its ability to behave as a reduction-oxidation cycler 6. Methylene blue is a cationic tricyclic compound that contains a central thiazine ring system and carries a positive charge at physiologic pH. These structural properties explains its high reduction potential, similar to oxygen. Unlike ephemeral antioxidants (e.g., ascorbic acid), methylene blue is capable of auto-oxidizing, which regenerates its ability to act as a reducing agent 7.
Moreover, methylene blue’s aromatic, tricyclic structure confers high lipophilicity and rapid blood-brain barrier penetration. Reports indicate that methylene blue readily enters the brain and accumulates in nervous tissue after parenteral or oral administration. (Molecules that dissolve more easily in hydrocarbons or oils than water have a better chance of making it into the brain. Because most amino acids are water soluble, for example, they would never enter the brain unless they were actively and specifically transported across the blood brain barrier).
Once in the mitochondria, methylene blue mimics the function endogenous electron carriers such as ubiquinone, by accepting and transferring electrons to oxygen or alternate electron acceptors. This redox behavior is not a recent discovery and has been appreciated for decades 8. (In order for the mitochondria- the energy powerhouses of the cell-to produce ATP, protons need to be pumped across the inner mitochondrial membrane. This proton pumping action is tightly coupled to a series of electron transfer reactions. Methylene blue supports the mitochondria by stepping in and accepting and donating electrons, improving the overall efficiency of the electron transport chain.)
The methylene blue dose-response curve is hormetic, meaning that its activity displays an inverted U-shape opposite effects at low doses vs. high doses 9. This effect may be a consequence of the fact that low mitochondrial methylene blue concentrations promote dimerization and reduction, whereas high concentrations favor oxidation and reaction with NADH and NADPH, the endogenous electron donors.
Hence, it is expected that low doses will be therapeutic and high doses pathologic. Methylene blue is proposed to enhance memory by amelioration of impaired mitochondrial oxidative metabolism in aged animals.
Early studies have noted that methylene blue increases oxygen consumption at the expense of carbohydrate metabolism. More recently it has been observed that methylene blue enhances oxidative metabolism via direct interaction with components of the mitochondrial respiratory chain, i.e., the protein complexes which are embedded in the inner mitochondrial membrane such as coenzyme Q, cytochrome c and complex I-IV.
Under native conditions, inner membrane proteins accept electrons from NADH and FADH2. Methylene blue supports the proton gradient by acting as a surrogate electron acceptor. Methylene blue’s ability to donate electrons to coenzyme Q and possibly cytochrome c and concomitantly increase oxygen consumptions is well-characterized 10.
MB and Neuroprotection: Evidence from Animal Models
|2006||MB rescues impairment in an animal model of optic neuropathy||Zhang X et al. Methylene blue prevents neurodegeneration caused by rotenone in the retina. Neurotox Res. 2006;9(1):47-57.|
|2010||High-dose MB rescues soluble tau burden and ameliorates cognitive impairment in tau transgenic mice||O’leary JC et al. Phenothiazine-mediated rescue of cognition in tau transgenic mice requires neuroprotection and reduced soluble tau burden. Mol Neurodegener. 2010;5:45.|
|Rojas JC et al. Striatal neuroprotection with methylene blue. Neuroscience. 2009;163(3):877-89.|
|2011||MB improves proteasome function and ameliorates AD-like pathology in vivo||Medina DX et al. Methylene blue reduces aβ levels and rescues early cognitive deficit by increasing proteasome activity. Brain Pathol. 2011;21(2):140-9.|
|2012||In a drosophila model of Huntington’s MB reduced neurodegeneration and reduced the disease phenotype in transgenic mice||Sontag EM et al. Methylene blue modulates huntingtin aggregation intermediates and is protective in Huntington’s disease models. J Neurosci. 2012;32(32):11109-19.|