1. α-Linolenic Acid—An Essential Nutraceutical
α-Linolenic acid (ALA) is an essential omega-3 polyunsaturated fatty acid (PUFA) that is found in green leaves, seed oil (flax), pumpkin seeds, beans and walnuts; flaxseeds are the richest source of ALA . ALA is an 18 carbon polyunsaturated fatty acid containing three double bonds at the 9, 12 and 15 positions. ALA plays an important role in brain function and protection as well as exhibiting anti-inflammatory and neuroplastic properties [2,3,4] and has a very wide safety margin [5,6]. ALA is a precursor of the long-chain PUFAs, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA).
Early work showed that administration of ALA resulted in an increase of the omega-3 PUFAs 7,10,1,3,16,19-docosapentaenoic acid and 4,7,10,13,16,19-docosahexaenoic acid in the brain . Humans and rats have the ability to metabolize ALA to form EPA and DHA, but the overall conversion appears to be limited in humans  and in rats . In a single male subject, a significant portion of ALA is converted into DHA covalently bonded to the 2-acyl position of phosphatidyl choline in plasma . However, the significance of this product in plasma in a single male subject as it applies to the pleotropic properties in brain mediated by the administration of ALA is unknown, raising the possibility that ALA exerts actions of its own. Addition of ALA to cells in culture where metabolism of ALA would not be expected to take place increased neuronal stem cell survival and reduced neuronal cell death in a model of N-methyl-d-aspartate (NMDA) receptor-mediated excitotoxicity . In weanling rats made deficient in essential fatty acids, administration of ALA resulted in a reduction in omega-6 polyunsaturated fatty acids such as arachidonic acid suggesting that omega-3 PUFAs exert their effect in brain by inhibiting the desaturation of dihomo-γ-linolenic acid to arachidonic acid. Although ALA was converted to docosahexaenoic acid, there were no differences in the brain levels of eicosapentaenoic acid, a metabolite of ALA . In contrast, administration of ALA in the form of perilla oil to a group of spontaneously hypertensive rats did not change the ratio of unsaturated to saturated phospholipids but there were marked differences in the proportion of omega-3 and omega-6 fatty acids compared with a group of rats administered safflower oil. The most notable difference was a decrease in the proportion of the omega-3 PUFA docosahexaenoate in phospholipids and an increase in the omega-6 PUFAs docosatetraenoic and docosapentaenoic acids compared with the perilla oil group of animals. Importantly, the correct response ratios were higher in the perilla oil group (high ALA) of animals compared with their safflower counterparts in a learning discrimination task . Administration of sunflower oil to rats which is low in ALA resulted in changes at the cellular level. For example, the sodium-ATPase activity in neuronal membranes was reduced by 40% whereas the activity of the 5’ nuclease enzyme was reduced by 20%; these changes were associated with significant learning impairment and neurons were more sensitive to injection of a neurotoxin . These cellular changes may in part be due to alterations in the fluidity within the plasma membrane. ALA has been shown to increase membrane fluidity which may maintain or restore membrane function in undamaged and damaged cells respectively . It has been suggested that the high ratio of omega-6/omega-3 PUFA may be involved in the pathogenesis of many diseases, including cardiovascular disease, cancer inflammatory and automimmune disorders . While many believe that a balance between omega-6/omega-3 PUFA may be important, the “ratio theory” remains controversial. Taken together, ALA has been shown to undergo conversion to docosapentaenoic and docosahexaenoic acids in brain but whether the conversion products are required for the actions of ALA is currently unknown.
2. ALA Protects against Animal Models of NMDA Receptor-Mediated Excitotoxicity
Glutamate is the major excitatory neurotransmitter in brain. Paradoxically, the pathophysiology of hypoxic-ischemic neuronal damage in acute and chronic neurodegenerative disorders involves glutamate. The glutamate receptor subtype N-methyl-d-asparate (NMDA) plays a major role in neuronal damage [15,16,17,18] and references therein).
In a well-established model of epilepsy induced by kainic acid, ALA treatment, but not other PUFAs or saturated fatty acids, was able to almost completely abolish neuronal cell death in the hippocampal CA1 and CA3 subfields. While other PUFAs exerted neuroprotective efficacy in vivo, ALA resulted in the most efficacious and reproducible effect . Surprisingly, the intravenous administration of ALA (500 nmol/kg) significantly increased the hippocampal levels of activated nuclear factor kappaB (NF-κB), a transcription factor, in a time- and concentration-dependent manner [20,21,22]. Furthermore, the increase in activated NF-κB levels in neurons played an essential role in mediating neuroprotection induced by ALA in vivo  and by subtoxic concentrations of NMDA against NMDA receptor-mediated excitotoxicity in vitro . It has been suggested that NF-κB is involved in neuronal plasticity in addition to its well-known role in inflammatory responses [23,24,25,26]. ALA was demonstrated to be neuroprotective in other models of hypoxic-ischemic neuronal injury [27,28,29,30].
3. Organophosphate (OP) Nerve Agent-Induced Excitotoxicity and the Limited Availability of Neuroprotective Therapies
Organophosphate (OP) nerve agents are some of the most deadly toxins known to man. The G series class of OP nerve agents includes soman, sarin, cyclosarin, tabun and VX. These agents penetrate the human body through skin, inhalation, and via the bloodstream. The rapidity of symptom onset depends upon the route of nerve agent exposure. Nerve agents inhibit acetylcholinesterase (AChE) quickly and completely and little to no spontaneous reactivation of the enzyme occurs following exposure to sarin, cyclosarin or soman. In the case of severe nerve agent exposure, absorption into the bloodstream occurs quickly and death occurs within minutes of the development of the cholinergic crisis secondary to respiratory and cardiovascular collapse. Absorption of a volatile nerve agent through the skin results in a more deliberate uptake and accumulation of nerve agent in the bloodstream leading to a slower cholinergic crisis .
There has been a disturbing resurgence in OP nerve agent use around the world against military and civilian populations by terrorist groups and organizations. The release of sarin in Matsumoto and in the Tokyo subway by a terrorist organization led to the intoxication of thousands of people and nineteen deaths [32,33,34]. The latest incident was in Syria where more than one thousand people, including 426 children, died in the aftermath of sarin deployment last year . Current therapy against exposure to nerve agents targets selective areas within the body to promote overall survival. Atropine, a muscarinic antagonist, reduces the attended copious secretions, bradycardia and gastrointestinal effects. Pralidoxime (2-PAM), an oxime, reactivates acetylcholinesterase molecules that have not undergone aging. After the phosphate moiety on OP nerve agent binds to the serine residue within the active site of AChE to form an ester, a process known as aging occurs whereby there is an internal dealkylation reaction leading to an OP nerve agent-acetylcholinesterase bond that cannot be reactivated by an oxime . The rate of aging is variable, depending on the toxicity of the nerve agent. Acetylcholinesterase will “age” in only two minutes after binding to soman .
The underlying mechanism of OP-induced toxicity is the inhibition of AChE which in turn leads to the excessive accumulation of the excitatory neurotransmitter acetylcholine (ACH) within synapses (the cholinergic phase) resulting in a plethora of signs and symptoms including Status epilepticus; overactivation of muscarinic receptors results in the generation of seizures and Status epilepticus [38,39,40,41,42,43]. Overstimulation of muscarinic receptors by the excessive synaptic levels of acetylcholine and ischemia secondary to the generalized seizures increases the release of glutamate [44,45,46] and γ-aminobutyric acid [GABA] [47,48] disrupting the balance between excitatory and inhibitory input resulting in Status epilepticus. The glutamate phase involves glutamate receptors which in turn participate in the propagation and maintenance of nerve agent-induced seizures; the N-methyl-d-aspartate (NMDA) glutamate receptor subtype plays a major role in excitotoxic-mediated neuronal cell death in vulnerable brain regions [49,50,51,52].
Prolonged seizures result in brain region-specific neuropathology leading to long-term cognitive and behavioral deficits in animals; long-term cognitive and behavioral deficits have been demonstrated in human survivors . The most profound neuropathology occurs in the amygdala followed by the hippocampus and piriform cortex [21,41]. In fact, the neuropathology can be observed in vulnerable brain regions even when seizures were stopped by benzodiazepines after five minutes . This result alone indicates the danger of exposure to OP nerve agents in survivors. Thus, diazepam, a benzodiazepine, stops/attenuates seizures, but does not prevent the neuropathology .
Cognitive and behavioral impairment have been observed years after exposure to OP nerve agents. For example, on the day of the Tokyo subway attack, 5500 individuals that were exposed to sarin gas were evaluated by hospitals. While most complained about minor symptoms, 1000 patients had moderate and 50 patients had severe signs of cholinergic crisis, respectively, associated with low plasma cholinesterase activity and there were 12 deaths . Forty-five percent of victims from the Tokyo subway attack that responded to a survey continued to exhibit symptoms one year after the incident in one study . Post-traumatic stress disorder (PTSD), anxiety, depression, lack of concentration, memory and cognitive deficits, impairment in motor function and coordination as well as structural changes in the right insular cortex and temporal cortex, left hippocampus and loss of white matter in the left temporal area near the insular region on brain MRI  develop months to years after exposure to OP nerve agents as well as OP insecticides [56,57,58,59,60,61,62,63]. Importantly, depression and memory impairment are serious and prominent features of exposure to OP nerve agents as well as pesticides [56,61,62,64,65,66]. While depression correlated with PTSD in those exposed to sarin in the Tokyo subway terrorist attack , depression is also observed in individuals exposed to OP pesticides [60,61] as well as those individuals exposed to sarin where red blood cell cholinesterase level is between 10% and 40% of control  raising the possibility that a cultural difference in Japanese people may have contributed to the depressive symptoms associated with PTSD . Alterations on brain MRI in white as well as gray matter were also observed in Gulf War veterans exposed to OP nerve agents at Khamisiyah [67,68].
Cognitive and behavioral deficits have also been observed in animal models of OP nerve agents. Learning and memory impairments were demonstrated in rodents exposed to OP nerve agents [69,70,71,72,73] including deficits in the Morris water maze and passive avoidance test which are hippocampal-dependent memory tasks [74,75,76]. Deficits in fear-based learning were also shown to be present after exposure to OP nerve agents [77,78]. PTSD and anxiety were demonstrated in rodents exposed to OP nerve agents [78,79,80]. For the first time, our group showed an increased immobility time on the Porsolt forced swim test indicative of a depressed-like state after OP nerve agent exposure . Altogether, the cognitive and behavioral deficits observed in animal models of OP nerve agents replicate the cognitive and behavioral impairment observed in humans that survived exposure to OP nerve agents.
The capability of OP nerve agents to cause mass casualties, the long-term morbidity of individual survivors after exposure to OP nerve agents and the modestly effective antidotal drugs provide the underpinnings for the development of new and more efficacious therapies.
Exposure to an OP nerve agent results in a plethora of signs and symptoms indicative of a cholinergic crisis in the peripheral and central nervous system (Figure 1) . The rapid increase of acetylcholine that occurs early in soman poisoning is known as the cholinergic phase. During this phase seizures can be blocked with muscarinic receptor antagonists when given immediately after nerve agent exposure. If not controlled, continued seizure activity recruits glutamate and possibly other neurotransmitters to propagate and maintain seizures [45,46,81]. This is the transitional phase with modulation of cholinergic/non-cholinergic systems. During this phase which is referred to as the glutamate phase, seizures cannot be stopped with muscarinic receptor antagonists. Moreover, glutamate can further stimulate the release of acetylcholine contributing to maintenance of the seizures and central nervous system (CNS) neurotoxicity [49,51]. Status epilepticus triggers a cascade of effects (overactivation of inotropic glutamate receptors, cytotoxicity, ion imbalance—Ca2+ influx—and inflammation) leading to hypoxic-ischemic injury and NMDA receptor-mediated excitotoxicity, the major contributor to neuronal death after OP poisoning.
Caramiphen is an anticholinergic drug used in the treatment of Parkinson's disease. In combination with phenylpropanolamine it is used as a cough suppressant and nasal decongestant to treat symptoms associated with respiratory illnesses such as cold, allergies, hay fever, and sinusitis.