Transcript – Using Raman Spectroscopy for the Detection of Opioid Use Disorders

Opioid use disorder is a serious and relapsing mental health disease. In humans, both personality and social environment were suggested to influence the development of opioid use disorder. Similarly, we demonstrated using rodent models that social environment and individual sociability levels influence the response to opioids. Mice can be tested and divided based on their sociability levels into socially avoiding and socially exploring groups. We demonstrated that socially exploring mice are more sensitive to the sensitizing effects of opioids. This suggests that the positive reward system is likely mediating abuse escalation in socially exploring individuals. In contrast, socially avoiding mice are more sensitive to opioids effects on stress and pain. This suggests that negative reinforcement is likely mediating abuse, escalation and socially avoiding individuals. Although opioid use disorder is a serious and relapsing mental health disease, in contrast to other chronic disorders, there are no objective diagnostic tools for early detection or to monitor disease progression. Raman spectroscopy is used in other biomedical fields for both research and as a diagnostic tool. But it is underutilized for the study of opioid use disorder. Raman scattering is the inelastic scattering of photons by matter. Transfer of energy from a photon emitted by a monochromatic light source to molecules results in the gain of vibrational energy. As such, the nondestructive technique Raman spectroscopy can be used to probe the vibrations within a molecule of interest. The Raman scattered photons can be measured as a function of their frequency difference compared to the incident photons. This is most commonly referred to as the Raman shift. Every molecule contains a unique set of vibrational modes, enabling Raman spectroscopy to discriminate between species, which exhibit structural similarity acutely.

The use of Vermont as an analytical tool extends beyond chemical analysis into complex biological interrogation. As such, it has been readily applied to probe disease states in clinical samples. The benefits of Vermont spectroscopy over other vibrational spectroscopic techniques include its nondestructive nature, minimal sample preparation, rapid spectral acquisitions, highly specific chemical and biological fingerprinting, and a lack of water interference. Raman Spectra were collected using a Wasatch Photonics Raman probe system equipped with a 785 nanometer laser at 115.6 milliwatts power. The focal distance of the laser via the probe is one point one centimeters delivered to give a laser resolution of 50 micrometers. S. mice brains were placed directly under Raman Laser and five spectra were collected in each brain region with five seconds integration time using enlightened spectroscopy software. The spectral data was analyzed on Matlab. We used multivariate analysis employing principle component analysis. Accordingly, here we demonstrate that using Raman spectroscopy, we can detect baseline differences between socially avoiding and socially exploring mice as well as in their responses to morphine. Socially avoiding and socially exploiting mice were treated with saline or morphine in the figure. Saline-treated socially avoiding mice are marked in green saline treated socially exploiting mice are marked in blue, morphine treated socially. Avoiding mice are marked in purple and morphine treated socially exploring mice are marked in red. Different brain areas were examined, including the amygdala nucleus accumbens, ventral tegmental area, prefrontal cortex and hippocampus for space considerations only.

The results from the amygdala and nucleus accumbens are shown to compare baseline differences between socially avoiding and socially exploring mice. We compared between saline treated socially avoiding mice and green and saline treated socially exploring mice in blue. These results are shown in the far column on the left hand to observe the effects of repeated morphine we compared between saline treated socially, avoiding mice in green and morphine treated socially, avoiding mice in purple as well as saline treated socially, exploring mice in blue and morphine, treated socially, exploring mice and read. These results are shown in the two middle columns to compare the differences in responses to morphine between socially avoiding and socially exploring mice. We compared morphine treated socially, avoiding mice in purple and morphine treated socially, exploring mice in red. These results are shown in the far column on the right end for each brain area. The upper graphs are the three scores plot and the lower graphs are two score plots. As can be seen, there is separation in PC one between SALINE socially avoiding and saline socially exploring. In both the amygdala and the nucleus accumbens, there is separation in PC one between saline socially avoiding and morphine socially avoiding in both the amygdala and the nucleus accumbens.

There is also separation in PC two between saline socially exploring and morphine socially exploring in the amygdala, but not in the nucleus accumbens. There is separation in PC one between morphine, socially avoiding and morphine, socially exploring only in the nucleus accumbens and not in the amygdala in the lower bottom right of the poster. We show the PC one loadings plot outlining the spectral bands responsible for discrimination of spectra taken from the amygdala for space consideration only one plot is presented. Visualization of the loadings can help deduce the nature of the molecular features responsible for cohort discrimination. The display loadings suggest alteration of phosphordiester bands, lipids/collagen and aimide II. In lay terminology, we observed molecular differences within the amygdala and the nucleus accumbens between socially avoiding and socially exploring mice. This is very exciting and indicates that humans’ individual differences in personality and social interactions could possibly be detected and assessed using Raman spectroscopy. Repeated exposure to opioids, altered brain molecular composition in both socially exploring and socially avoiding mice. However, differential responses were observed in the response to morphine and socially avoiding and socially exploring mice. That is morphine-induced similar changes in the amygdala of both socially avoiding and socially boring mice. However, morphine induced changes in nucleus accumbens were observed only in socially avoiding and not in socially exploring mice. These suggest that socially avoiding and socially exploring individuals have differential behavioral and molecular responses to opioids, and that treatment for opioid use disorder should be tailored to individuals based on their personalities and social interaction style. Thus, these methodologies are expected to provide insights into potential need to individualize the care for opioid use disorder patients, as well as to provide markers to be used for objective diagnosis of the development of the trajectory of opioid use disorder in humans.