Considering the brain’s high dependence on oxidative metabolism, it is not surprising that various cognitive problems have been seen in patients with mitochondrial encephalomyopathies, in addition to other neurological abnormalities. A large cohort study on adults with mitochondrial disorders revealed a global pattern of cognitive dysfunction in patients, and specific studies focused on MELAS patients also showed global deficiencies in neuropsychologic performance [3
]. The natural history study being conducted at Columbia University Medical Center has already demonstrated that MELAS patients manifest severe cognitive deficits including impaired reasoning, memory, language, attention, and visuo-spatial orientation [14
]. Despite this epidemiologically and clinically well-recognized association between cognitive and psychiatric problems and MELAS, the biological basis of these problems is not well understood. In this study, we explored the possible role of CB in the pathogenesis of cognitive dysfunction, and demonstrated a significant reduction of CB in the MELAS hippocampus by immunohistochemistry, western blot, and real-time RT-PCR. Various previous studies in aging [15
], Alzheimer’s disease (AD) [18
], and other neurodegenerative disorders [19
] have pointed out alterations of hippocampal CB expressions in human cognitive dysfunction. In our study, all four MELAS patients had shown memory and/or learning disability, and the reduced hippocampal CB expressions probably resulted in unbuffered increase of intracellular calcium in the region, and may account for the patients’ hippocampal dysfunctions. Notably, animal studies have related altered CB expressions to deficits of hippocampal long-term potentiation (LTP), a primary experimental model of memory formation in neuronal circuits [6
]. Thus, LTP deficits could be one neurophysiologic dysfunction causing memory/learning disability in the MELAS patients, similar to what has been proposed in Alzheimer’s disease [6
]. Recent studies on LTP support the notion that at least three forms of LTPs are mechanistically separated at synapses in the dentate gyrus and CA1 of the hippocampus, and each of them may play a different role in learning and memory processing [22
]. It is conceivable that the significant reduction of ATP in MELAS [25
] could interfere with phosphorylation of various enzymes critically involved in LTP induction, maintenance, and expression and result in clinical memory impairment appearing at a much earlier age than in patients with sporadic AD.
The precise mechanism of the CB reduction in causing hippocampal dysfunction in MELAS remains unclear. In this study, the degree of CB reduction did not correlate with regional mutation loads. Neither did it appear to correlate with regional neuronal loss: the histological examination of MELAS hippocampal sections did not reveal a neuronal loss comparable to the profound CB reduction detected at the protein and messenger levels. Considering that all of our patients had seizures, altered CB expressions might be ascribable, at least in part, to seizure activity, possibly via synaptodendritic injury in hippocampal circuits. This could be associated with axonal sprouting or altered GABA receptor expression among dentate granular neurons and hippocampal interneurons [26
]. Interestingly, our preliminary Western blot data show that CB is variably reduced not only in the hippocampus, but also in frontal, temporal, parietal, and occipital cerebral cortices of all patients (data not shown). This allows us to further postulate that loss of CB could be a global phenomenon rather than an isolated event in the intrinsic hippocampal circuits. However, on the basis of our data in MELAS autoptic brains, it is difficult to discriminate whether this cerebral cortical CB reduction is due to a functional alteration of the protein at the cellular level, or to a selective loss of CB-bearing neurons including inhibitory GABAergic neurons. This problem is due to the fact that all our patients had multiple cerebral infarcts, tissue loss, edema, or reactive gliosis associated with infarction, all of which might have influenced the regional neuronal density and/or neurotransmitter distribution.
Nonetheless, the significant reduction of CB, once it occurs, could easily place the regional neuronal circuit in a vicious cycle of malfunction, if not cause outright cell death. Given the mitochondrial ATP starvation in the MELAS brain, the voltage-dependent calcium ion channels of inhibitory neurons may work improperly, allowing a large influx of calcium into the cells [30
]. In addition, cells harboring the m.3243A>G mutation may not be able to sequester excessive calcium, due to a decrease in mitochondrial membrane potential [31
]. The already abnormal cellular calcium homeostasis would be further exacerbated by the depletion of calcium binding proteins. Moreover, cell surface receptors on interneurons and astrocytes that are activated by both exogenous and endogenous ATP [32
] may not be adequately activated in MELAS brains with faulty mitochondrial respiratory chain function, resulting in decreased synaptic inhibition of hippocampal circuits, and increased excitatory neuronal activities. Clinically, this could mean additional seizures in MELAS patients.
In conclusion, we demonstrated in this study a significant reduction of CB in the post mortem hippocampal tissues from MELAS patients. We believe that the depletion of CB is not merely due to the regional neuronal loss, and that it may be relevant to the compromised cognitive functions often observed in MELAS patients.