Traumatic brain injury
Experimental studies have now established a critical role for brain free magnesium concentration in a variety of pathological conditions of the central nervous system including traumatic brain injury, stroke, drug intoxication, headache, seizures and neurodegenerative diseases, amongst others . The first of these demonstrations occurred in traumatic brain injury using magnetic resonance spectroscopy to determine brain free magnesium levels . In those studies, brain free magnesium was shown to fall by approximately 50% within hours of the injury. Total magnesium concentration, as measured using atomic absorption spectrophotometry, also declined by a much smaller 15%, reflecting the buffering capacity of the total magnesium pool. Magnesium levels remained low for at least three to five days before returning to normal levels , with this period of decline thought to represent a period of increased vulnerability to subsequent injury. Notably, restoring brain magnesium levels back to baseline values after trauma, using various pharmacological approaches, improved outcome [4, 5]. These findings have now been reproduced in a variety of experimental brain injury models in several different animal species, and have been described in hundreds of papers published by different laboratories [1, 6]. It is therefore widely accepted that brain free magnesium concentration plays a critical role in the secondary injury cascade following acute brain injury, and this has been a paradigm shift in understanding the role of magnesium in brain function.
On the basis of these results in experimental studies, clinical trials examining the efficacy of magnesium treatment following traumatic brain injury soon followed. In a small study of severe, closed traumatic brain injury, Dhandapani et al. reported that parenteral magnesium administration reduced mortality, intraoperative brain swelling, and outcome at three months when the total dose was administered to patients within 24 h of the original brain injury . Notably, the patients all had an initial Glasgow Coma Scale score of between five and eight, which is a relatively narrow injury range aimed at achieving reasonable homogeneity amongst the patient group. In contrast, the larger cohort study by Temkin et al. administered sustained parenteral magnesium over a five day-period to a more heterogeneous injury group with an initial Glasgow Coma Scale score ranging between three and twelve . No benefit was recorded. Interestingly, the controls in the latter study also received magnesium if their serum level was below 0.75 mM, which was part of standard care and targeted at restoring serum magnesium levels back to normal levels. Thus, the study essentially compared restoring serum magnesium levels to normal after traumatic brain injury (which was the standard protocol in the experimental studies), versus administration of high levels of magnesium over a prolonged period of time. As in the experimental studies, high levels of magnesium were ineffective at improving outcome , and at times were deleterious.
Irrespective of the clinical trial methodology, questions were raised with respect to CNS penetration of magnesium salts when administered after a 24 h time period, given that 24 h is considered to be the time period in which the blood brain barrier is maximally open after traumatic brain injury, and thus permissive for magnesium entry . Indeed, several studies have suggested that parenteral magnesium administration does not result in an increase in CSF magnesium under conditions of severe traumatic brain injury with intracranial hypertension and where the blood brain barrier is thought to be intact [11, 12]. Thus, strategies need to be developed that will facilitate CNS penetration of magnesium under these conditions.
In an effort to address this lack of CNS penetration by magnesium salts, magnesium sulphate was combined with polyethylene glycol, which has been used to enhance transmembrane transport of various substances , with the combination shown to improve outcome after spinal cord injury in rats . Similar protective effects were subsequently demonstrated in traumatic spinal cord injury using magnesium chloride and polyethylene glycol combined, with these studies also establishing an optimal dose and the therapeutic window . The success of these experimental studies in spinal cord injury inspired a similar set of studies to be undertaken in traumatic brain injury . Specifically, magnesium chloride in polyethylene glycol was shown to be as effective on both motor and cognitive outcome after traumatic brain injury as an optimal dose of magnesium chloride alone, even when the magnesium concentration in the polyethylene glycol mixture was reduced to 10% of the optimal dose. Indeed, with respect to motor outcome (figure 1) and brain water content (edema), the reduced magnesium dose in polyethylene glycol was superior to the optimal dose of magnesium alone. Histopathological examination of the injured brains demonstrated reduced dark cell change and preserved cellular architecture on hematoxylin and eosin-stained hippocampal sections, suggesting that the combination of magnesium in polyethylene glycol had direct neuroprotective actions. Thus, it would seem that the combination of magnesium in polyethylene glycol enhances CNS penetration of the salt, even at reduced concentrations, thus avoiding potentially negative side effects of high peripheral magnesium concentrations while still achieving central concentrations sufficient to be neuroprotective.
It has been known for some time that chronic exposure to alcohol causes magnesium deficiency and reduces brain free magnesium concentrations [17, 18]. Acute alcohol exposure has also been shown to reduce brain free magnesium concentration and has been implicated in predisposing affected individuals to headache and potentially stroke . With the growing incidence of alcohol-associated violence in some western countries, and the legal implications of “one-punch” head injuries, our laboratory has investigated the association between alcohol intoxication, brain free magnesium concentration, and outcome following traumatic brain injury . Our results demonstrated that a moderate traumatic brain injury rapidly reduced brain free magnesium concentration in both acute and chronic alcohol-intoxicated animals to levels lower than in non-intoxicated animals. Moreover, the levels of magnesium remained lower for a longer period of time than in non-intoxicated animals, thus potentially exposing the brain to greater secondary injury. Given the demonstrated association between low magnesium levels and exacerbation of injury following head trauma , we therefore postulated that alcohol intoxication sensitizes an individual to a worsened outcome should a traumatic brain injury ensue. This is certainly consistent with recent literature reports that demonstrate an augmentation of concussive brain injury by alcohol . Thus, reduced brain magnesium concentration induced by various ingested drugs becomes an important determinant of outcome in a concussive event that might not normally be expected to result in a catastrophic outcome.
Stroke is another form of acute brain injury where magnesium has been implicated as an important determinant of outcome . Indeed, a number of studies have demonstrated that magnesium administration after stroke reduces infarct volume and improves outcome . However, the intravenous magnesium efficacy in stroke (IMAGES) trial clearly demonstrated no overall beneficial effect of magnesium in stroke , albeit there was some suggestion of efficacy in lacunar strokes, an observation that was subsequently confirmed in a later analysis . Given that the experimental animal studies typically used early administration of magnesium to demonstrate a neuroprotective effect, a clinical trial was subsequently conducted to examine the efficacy of early magnesium administration on stroke outcome . Again, the trial failed, with magnesium administered within two h of stroke onset showing no improvement in disability outcomes at 90 days.
The question remains as to what might account for these marked differences between traumatic brain injury and stroke outcomes after magnesium administration? One fundamental difference between the two injuries is that stroke results in complete energy depletion, that is, a total loss of ATP. In contrast, while traumatic brain injury does result in hypometabolism and compromised energy production, it typically does not result in ATP depletion. One of the fundamental properties of magnesium is as an essential cofactor in all energy producing and consuming reactions, a property that would facilitate a cell's recovery from injury. Of course, when ATP has been depleted, magnesium would be ineffective at promoting cell recovery mechanisms at the metabolic level. It may therefore be that under conditions of energy depletion as occurs in stroke, we are being too optimistic in believing that magnesium treatment will be of significant benefit. Indeed, the beneficial effects of magnesium would be limited to those mechanisms that do not require energy metabolism. In contrast, in injuries such as traumatic brain injury, where there are still significant reserves of ATP present after the insult, magnesium will be beneficial through both ATP dependent and non-dependent pathways. In the case of lacunar stroke, where beneficial effects of magnesium were observed in a clinical situation, it is likely that collateral flow to the white matter was able to preserve some energy metabolism and thus provide ATP for repair processes.
There have been various reports of magnesium depletion in Alzheimer's disease, including in postmortem brains, cerebrospinal fluid, hair, and even of free magnesium in the blood serum . However, simple administration of magnesium salts has not been effective in either attenuating disease progression or in reversing the symptoms of the disease. As alluded to in the section on traumatic brain injury, an intact blood brain barrier may prevent serum magnesium from readily crossing into the brain, thus prevent therapeutic levels from being achieved centrally. A research team led by Guosong Liu has approached this dilemma in an animal model of Alzheimer's disease by comparing different salts of magnesium and examining their effects on synaptic density and cognitive deficits . This group reported that the threonate form of the magnesium salt showed superior central penetration to other forms of magnesium salts, reduced the presence of amyloid-beta plaques, significantly increased synaptic density (p<0.001), and improved cognitive function. Moreover, the synaptic activity in treated mice was greater than in mice that were not treated with the magnesium compound.
Similar beneficial effects of magnesium threonate have also been reported on neural stem cell proliferation in the rodent hippocampus . Specifically, these authors were able to demonstrate in both young and aged animals that either short-term or long-term dietary supplementation with magnesium threonate increased the numbers of hippocampal neural stem cells, thus attenuating the decline in these cells that occur with age. Moreover, the beneficial effects of magnesium seemed to be mediated through enhanced energy metabolism downstream from the mitochondria . Given that short-term memory is affected early in Alzheimer's disease, any increase in neurogenesis in the hippocampus would likely contribute to improved cognitive performance of affected individuals.
Finally, the observation that magnesium reduced BACE1 expression in the transgenic animals  was particularly noteworthy given previous studies. BACE1 is a key enzyme in the amyloidogenic pathway that cleaves amyloid precursor protein (APP) to produce the neurotoxic amyloid beta. Alternatively, APP can also be cleaved by α-secretase, resulting in soluble amyloid precursor protein alpha, a protein that has been shown to increase synaptogenesis and improve functional performance . Whether the cleavage of APP occurs through the amyloidogenic BACE1 pathway or the non-amyloidogenic α-secretase pathway is dependent on the balance of positive and negative factors that influence neuronal survival and proliferation. For example, negative factors such as magnesium decline, inflammation, oxidative stress, hormone imbalance, and lack of nerve growth factors, amongst others, promote BACE1 activity and formation of amyloid beta. This along with other factors may contribute to the development of neurodegeneration. In contrast, the absence of these negative factors facilitates cleavage via the beneficial α-secretase pathway, thus, in part, preventing neurodegeneration. The previously published observation following traumatic brain injury that magnesium promotes non-amyloidogenic cleavage of APP  supports the view that restoration of magnesium homeostasis may thus be of benefit in Alzheimer's disease.
Considerably more research is required to examine whether the threonate compound has superior blood brain barrier penetration to other magnesium salts, and how this enhanced penetration is facilitated. Nonetheless, it is clear that facilitation of central penetration of magnesium salts is having beneficial effects on a variety of pathological secondary injury factors, many of which are dependent on the effects of magnesium on energy metabolism.
One major recent advance in understanding central nervous system disease has been the realisation that inflammation may play a major role in the pathology underlying many neurological conditions. Not only is inflammation involved in acute conditions such as traumatic brain injury and stroke, it has also been implicated in chronic and neurodegenerative conditions such as Alzheimer's disease and Parkinson's disease . Given some key differences exist between neuroinflammation and peripheral inflammation, in part because of the blood-brain barrier, the classical anti-inflammatory agents, such as steroids and non-steroidal anti-inflammatories, have not proven useful in most instances of neuroinflammation. However, antagonists of neurogenic inflammation, and particularly those targeting the action of substance P, have proven particularly effective .
Neurogenic inflammation is a neurally-elicited inflammatory response characterised by vasodilation, plasma exudation, tissue swelling and mast cell degranulation . It is mediated by the release of neuropeptides including substance P and calcitonin gene-related peptide from capsaicin-sensitive, sensory nerve fibres in response to factors such as histamine, serotonin, leukotrienes, pH change, temperature and mechanical stimulation. Cerebral blood vessels have a dense supply of sensory nerve fibres and have been shown to exhibit characteristics of neurogenic inflammation in response to various stressors .
Magnesium's role in classical inflammation and its ability to attenuate inflammatory responses is well known and has been described in detail elsewhere [37, 38]. Its role in neurogenic inflammation however, is less clear although recent reports suggest that it may also play a role in this form of neuroinflammation. Weglicki and colleagues were the first to demonstrate that hypomagnesia increased substance P release , thus initiating neurogenic inflammation and subsequent classical inflammation. Magnesium administration also attenuated substance P release, and accordingly downregulated neurogenic inflammation . In traumatic brain injury, we have demonstrated that there is an early release of substance P after injury , and that this release is associated with the decline in brain magnesium concentration. Indeed, inhibition of substance P release with capsaicin pre-treatment significantly attenuated the post-traumatic decline in brain intracellular free magnesium concentration . Moreover, treatment with a substance P antagonist increased brain intracellular free magnesium concentration after traumatic brain injury . Thus, there is a clear association between neurogenic inflammation and magnesium, although more research is required to characterise the relationship between magnesium treatment and ongoing neurogenic inflammation.
While a potential neuroprotective role for magnesium in neurological disease has been appreciated for almost three decades, translation to the clinical arena has proven particularly elusive. In part, this has been due to an inadequate understanding of magnesium transport across the blood brain barrier and how this can be facilitated. Various approaches to increase central penetration are now under investigation, with beneficial effects being reported in both acute and chronic pathological conditions. Many of these conditions feature neuroinflammation as an injury factor, and the characterisation of magnesium's effects on neuroinflammation will pave the way to a more targeted approach to magnesium therapy in diseases of the central nervous system.
Financial support: none. Conflict of interest: none.
* Presented at The XIV International Magnesium Symposium, Magnesium and Health, Rome, Italy, June 23-24, 2016