Significance: Decline in NAD ⁺ availability and abnormal NAD ⁺ /NADH redox state are tightly linked to age-related metabolic diseases and neurodegenerative disorders. To better understand the roles of NAD metabolism and redox state in health and disease, it is important to assess the intracellular NAD and redox state in situ. We report herein the first in vivo NAD assay, to our knowledge, that is capable of noninvasively and simultaneously measuring intracellular NAD ⁺ and NADH concentrations and NAD ⁺ /NADH ratio in the human brain and detecting the age-dependent changes in NAD contents and redox state associated with the normal aging. This method can potentially be applied to study various metabolic and neurodegenerative disorders by monitoring the NAD and redox state changes associated with disease progression or treatment in human patients.

A majority of ATP in the brain is formed in the mitochondria through oxidative phosphorylation of ADP with the F ₁ F ₀ -ATP (ATPase) enzyme. This ATP production rate plays central roles in brain bioenergetics, function and neurodegeneration. In vivo ³¹ P magnetic resonance spectroscopy combined with magnetization transfer (MT) is the sole approach able to noninvasively determine this ATP metabolic rate via measuring the forward ATPase reaction flux ( F _f,ATPase ). However, previous studies indicate lack of quantitative agreement between F _f,ATPase and oxidative metabolic rate in heart and liver. In contrast, recent work has shown that F _f,ATPase might reflect oxidative phosphorylation rate in resting human brains. We have conducted an animal study, using rats under varied brain activity levels from light anesthesia to isoelectric state, to examine whether the in vivo ³¹ P MT approach is suitable for measuring the oxidative phosphorylation rate and its change associated with varied brain activity. Our results conclude that the measured F _f,ATPase reflects the oxidative phosphorylation rate in resting rat brains, that this flux is tightly correlated to the change of energy demand under varied brain activity levels, and that a significant amount of ATP energy is required for “housekeeping” under the isoelectric state. These findings reveal distinguishable characteristics of ATP metabolism between the brain and heart, and they highlight the importance of in vivo ³¹ P MT approach to potentially provide a unique and powerful neuroimaging modality for noninvasively studying the cerebral ATP metabolic network and its central role in bioenergetics associated with brain function, activation, and diseases.

Spontaneous hemodynamic signals fluctuate coherently within many resting-brain functional networks not only in awake humans and lightly anesthetized primates but also in animals under deep anesthesia characterized by burst–suppression electroencephalogram (EEG) activity and unconsciousness. To understand the neural origin of spontaneous hemodynamic fluctuations under such a deep anesthesia state, epidural EEG and cerebral blood flow (CBF) were simultaneously recorded from the bilateral somatosensory cortical regions of rats with isoflurane-induced burst–suppression EEG activity. Strong neurovascular coupling was observed between spontaneous EEG “bursts” and CBF “bumps,” both of which were also highly synchronized across the 2 hemispheres. Functional magnetic resonance imaging (fMRI) was used to image spontaneous blood oxygen level–dependent (BOLD) signals under the same anesthesia conditions and showed similar BOLD “bumps” and dependence on anesthesia depth as the CBF signals. The spatiotemporal BOLD correlations indicate a strong but less-specific coherent network covering a wide range of cortical regions. The overall findings reveal that the spontaneous CBF/BOLD fluctuations under unconscious burst–suppression anesthesia conditions originate mainly from underlying neural activity. They provide insights into the neurophysiological basis for the use of BOLD- and CBF-based fMRI signals for noninvasively imaging spontaneous and synchronous brain activity under various brain states.