Fluorescence Sensing Identifies Cancer Indicators in the Brain

Researchers at the University Hospital Münster and Paris-Saclay University established a method to correct for artifacts caused by changes in blood flow, and recorded cell-specific lactate levels in rat brains using fluorescence signals from fiber-based fluorescence resonance energy transfer (FRET) sensors. While FRET is known to be useful for studying neurophysiology through microscopy, its use for in vivo studies is limited by artifacts that appear in the FRET recordings.

Lactate levels in the body can be an indicator for cancer and other diseases. Current methods for monitoring lactate do not have enough temporal resolution to detect real-time changes in lactate levels, especially in the brain, where lactate concentrations are low.

The researchers used Laconic, a lactate sensor, and Twitch-2B, a calcium sensor, to acquire FRET recordings of lactate and calcium levels in rats. They used functional MRI (fMRI) and pharmacological MRI (phMRI) signals to identify and correct hemodynamic artifacts in the recordings.

The research investigated whether fiber-based FRET sensors could successfully detect changes in lactate levels in vivo. The work also sought to determine how the sensors would respond to changes in blood flow and blood oxygen levels in the brain.

Laconic, a lactate sensor with high temporal resolution, contains two fluorescent chemical compounds: a donor compound and an acceptor compound. These compounds, known as fluorophores, emit light of different wavelengths and are separated by a molecule that interacts with lactate.

When the Laconic sensor was irradiated with light, the donor fluorophore absorbed the light and transferred it to the acceptor. The intensity of the emitted light depends on the energy transfer and the distance between the two fluorophores. When lactate interacted with the molecule separating the fluorophores, the distance between the donor and acceptor changed, which affected the intensity of the emission. The sensor detected the presence of lactate by observing the intensity of the emitted light. This information can be used with an fMRI scan to map cell-specific lactate levels and link them to the blood flow in the brain, the researchers said.

In vivo, fiber-based metabolite detection in the rodent brain using FRET sensors is feasible. For detection of lactate levels, correction of hemodynamic artifacts is required, which can be achieved by using simultaneously or separately acquired fMRI signals. Courtesy of H. Lambers/University of Münster.

The researchers genetically encoded the Laconic and Twitch-2B sensors to be expressed in the rats’ sensory cortices (the region of the brain that receives sensory information). To acquire the fluorescence signals, the team implanted an optical fiber just above the sensors to guide the light going in and out of the brain. After intravenously injecting the rat with lactate and stimulating the rat’s sensory cortex, the researchers measured the fluorescence signals expressed by the sensor implants.

Both the lactate and calcium sensors responded to changes in brain activity. The lactate sensor detected the increase in lactate concentration introduced by the injection. However, when the researchers compared the time courses of fluorescence, fMRI, and cerebral blood volume signals, which were recorded upon sensory stimulation and the lactate injection, they identified prominent effects of hemodynamic artifacts. Artifacts were present in the ratios of fluorescence signals that corresponded to the lactate and the calcium signals for Laconic and Twitch-2B, respectively.

Because MR contrast is sensitive to hemodynamic changes, the researchers used MR-derived parameters to correct the hemodynamic artifacts in the fluorescence recordings. They used a correction algorithm that removed the artifacts for both simultaneous and separate measurements of fluorescence and MRI signals.

The MR-based correction algorithm enabled the researchers to detect lactate and calcium changes during sensory stimulation and intravenous lactate injections. The team validated the results of its measurements using local field potential recordings, magnetic resonance spectroscopy, and blood analyses.

Results indicated that fluorescence-based sensing could be used to measure cell-specific lactate level changes in living organisms, provided the signals are corrected for artifacts. The researchers’ approach could be used to provide additional information leading to the detection of cancer and other pathologies such as inflammatory and autoimmune disorders.

The research was published in Neurophotonics (www.doi.org/10.1117/1.NPh.9.3.032212).