Levels of neurotransmitters such as ATP in the brain can indicate brain health and neurodegenerative disorders such as Alzheimer’s disease. However, the protective blood-brain barrier (BBB) makes it difficult to efficiently deliver to the brain aptamer-based fluorescent sensors that can detect these small molecules. Researchers at the University of Texas at Austin Department of Chemistry have now developed a method that uses brain-cell derived exosomes to package ATP-responsive aptamer sensors for easy passage across the BBB in mouse models of Alzheimer’s disease, allowing for improved brain imaging. The team suggests that with further development the technology could help advance Alzheimer’s disease diagnosis and treatment.
Yi Lu, PhD, is corresponding author of the team’s published paper in ACS Central Science, which is titled, “Delivering DNA Aptamers Across the Blood−Brain Barrier Reveals Heterogeneous Decreased ATP in Different Brain Regions of Alzheimer’s Disease Mouse Models.” In their paper, the investigators stated, “Since DNA aptamers have been obtained for many other targets, the method developed in this work can be applied to deliver sensors across the BBB to image a wide range of other brain-related metabolites.”
Metabolites such as ATP play essential roles in neurobiology by serving as neurotransmitters and maintaining ion gradient flux, the authors wrote. It is common for neurotransmitter levels to decrease with age, but low levels of the neurotransmitter ATP can be an indication of Alzheimer’s disease. “Therefore, detecting the varying concentrations and cellular locations of these targets in the brain may help with understanding their roles in promoting brain health and preventing neurodegenerative diseases,” the team commented. However, they pointed out, “… in contrast to progress made in imaging macromolecules, such as nucleic acids and proteins, the selective detection of small molecule metabolites in the brain has been limited.”
One approach for measuring the location and amount of ATP in the brain uses fluorescent sensors based on pieces of DNA called aptamers, which light up when they bind to a target molecule. “… many DNA aptamer sensors for metabolites implicated in brain function, such as ATP, glucose, and dopamine, have already been developed,” the team commented.
Methods for delivering these sensors from the bloodstream to the brain have been developed, but most contain synthetic components that can’t easily cross the BBB. “The BBB is a specialized layer of endothelial cells that preserves brain homeostasis and prevents brain uptake of nonessential small molecules and macromolecules due to the selectivity of tight junctions between the endothelial cells,” the investigators explained. While a number of BBB-penetrable delivery techniques, such as cell-penetrating peptides, liposomes, microbubbles, and aptamer-based targeting ligands, have been tested, their application in vivo has been limited, partly because they use synthetic components that aren’t native to the brain and/or are not recognized by BBB transporters.
To overcome such limitations and develop a delivery system for sensors for live brain imaging, Lu and colleagues encapsulated their ATP aptamer sensor in brain-cell-derived microscopic vesicles called exosomes. “Exosomes derived from brain cells contain proteins and other macromolecules on their surface that can be inherently recognized by the BBB, imparting higher specificity for the BBB and increasing their biocompatibility and delivery efficiency compared with synthetic delivery vehicles,” they pointed out.
The scientists tested the new sensor delivery system in lab models of the BBB and in mouse models of Alzheimer’s disease. The BBB laboratory model consisted of a layer of endothelial cells on top of a solution containing brain cells. The researchers found that the sensor-loaded exosomes were nearly four times more efficient than conventional sensor delivery systems at passing through the endothelial barrier and releasing the fluorescent sensor into the brain cells. This was confirmed by measuring the observed level of ATP-binding-induced fluorescence. “We found evidence that the system uses recycling endosomes to transfer the sensors between the delivered exosomes and native endosomes, resulting in its high delivery efficiency,” they stated.
Next, Lu’s team injected mouse models of Alzheimer’s disease with either the sensor-loaded exosomes or free-floating unloaded sensors. By measuring fluorescence signals in the mice, the researchers found that the free-floating sensors stayed mainly in the blood, liver, kidneys, and lungs, while sensors delivered via exosomes accumulated in the brain.
In mouse models of Alzheimer’s disease, the exosome-delivered sensors identified the location and concentration of ATP in different regions of the brain. Specifically, they observed low levels of ATP in the hippocampus, cortex, and subiculum regions of the brain, which are indicative of the disease. “While previous Alzheimer’s research studies have also found decreased levels of both targets in whole brain tissue and patient serum, we were able to provide spatial resolution in live tissue, demonstrating that the decreased levels of ATP were not uniform throughout the brain,” they stated. “These results demonstrated the importance of the brain region type to contributing to disease pathology.” The researchers say that their exosome-loaded ATP-reactive sensors show promise for noninvasive live brain imaging and could be developed further to create sensors for a range of clinically relevant neurotransmitters. “Our method can be expanded to a wide variety of other aptamer sensors developed for other metabolites,” they noted.
