Sleep deficiency is a common challenge during pregnancy. Lack of sleep puts pregnant individuals and their fetuses at risk for poorer health outcomes. Pregnancy symptoms are known to be one of the main obstacles that prevent pregnant individuals from getting quality sleep. Mindfulness is a therapeutic practice where individuals focus on their feelings, thoughts, and bodily sensations in the present moment. Mindfulness meditation targets emotional and cognitive reactivity, which is a common pathway for developing depression and insomnia symptoms. Mindfulness-based interventions are known to help with sleep disorders, but little research has been conducted in regards to the possible relationship between mindfulness and sleep during pregnancy. A cross-sectional study was done to explore the associations amongst sleep hygiene, mood, and mindfulness. The aim of this study is to evaluate the relationship between pregnancy symptoms and morning restfulness in pregnant individuals with insomnia prior to the mindfulness intervention. We are examining the relationships among daily sleep habits, pregnancy symptoms, and quality of sleep in sleep diaries of pregnant individuals with insomnia. Preliminary findings show stronger relations between symptoms of worry and sickness to poorer sleep quality. These preliminary results will better inform mindfulness-intervention design specific to pregnant individuals with insomnia. 

The transient receptor potential vanilloid-1 (TRPV1) ion channel is a polymodal receptor expressed in sensory neurons that contributes to inflammatory pain sensation. Interestingly, the receptor channel is modulated by specific phosphoinositide lipid molecules in the plasma membrane. During inflammation, phosphoinositide 3-kinase (PI3K) generates the signaling lipid phosphatidylinositol (3,4,5)-triphosphate (PIP3). This increase in PIP3 levels leads to trafficking and fusion of vesicles containing TRPV1 with the plasma membrane. Previous research from the Gordon Lab revealed that increased PI3K activity is potentiated by TRPV1 during inflammation. However, the mechanism behind potentiation is undetermined. We hypothesize that stabilized association of PI3K with the plasma membrane by TRPV1 results in increased PI3K activity. To visualize the spatial localization of PI3K and its interaction with TRPV1, we employed a special optical technique called total internal reflection fluorescence microscopy (TIRFM). In TIRFM, selective excitation of fluorophores adjacent to the plasma membrane allows us to study translocation of proteins from the cytosol to the plasma membrane, or vice versa. We utilize TIRFM in conjunction with an opto-PI3K system to better understand PI3K signaling events during inflammation. We transiently transfected F-11 cells, model cells for dorsal root ganglion neurons that sense pain, with fluorescently labeled opto-PI3K system components and TRPV1 ion channels. Our fluorescence measurements show that, upon stimulation with 650 nm light, PI3K is retained at the plasma membrane through interaction with TRPV1. TRPV1 retention of PI3K is direct evidence for the hypothesized mechanism of potentiation through membrane localization. Studying PI3K activity in the prescence of TRPV1 will elucidate key properties of potentiation, such as affinity of the interaction, effects on PI3K activity, and substrate selectivity. These findings will aid in the development of inflammatory pain treatments that disrupt the interaction between PI3K and TRPV1.

Women fare significantly worse than men when using web-based alcohol interventions. These findings in the civilian population are mirrored among US veterans as evidenced by women veterans experiencing worse outcomes following completion of the VA’s web-based alcohol intervention “VetChange” compared to their male counterparts. Research is needed in order to inform adaptations to the “VetChange” web-based alcohol intervention that improve outcomes for women veterans. This project utilized a “think-aloud” methodology to collect data from women veterans with substance use disorder (SUD) and clinicians treating women veterans with SUD about their opinions of the current “VetChange” program. In accordance with “think-aloud” methodology, researchers observed participants’ use and impressions of VetChange during a period of unstructured access to the website, followed by a semi-structured qualitative interview to gather overall impressions of VetChange and what changes they recommend. Expected results include the ways in which the web-based intervention can improve two specific outcome measures, situational confidence and recovery-related coping behaviors, in order to adequately address women veteran’s SUD. The protocol and eventual data collection are of critical importance given that increasing numbers of women veterans are seeking SUD treatment and women veterans are dying at disproportionately younger ages than their civilian counterparts. Moreover, findings from this project can inform effective web-based interventions for women in the civilian population.

Underlying the mechanism of many biological processes are biomolecules called proteins. Proteins have dynamic 3D structures, fluctuating between an ensemble of different shapes (conformations), which often have varying physical and chemical properties. Thus, to understand how proteins function, we must understand their conformational dynamics: the kinetics and energetics associated with a protein’s conformational landscape. Here, we develop a novel combination of two spectroscopy techniques for probing protein conformational dynamics: single-molecule detection, and transition-metal-ion-fluorescence-resonance-energy-transfer (tmFRET). tmFRET is the distance-dependent transfer of energy between a donor fluorophore and an acceptor transition metal ion, and this phenomenon has previously been used to detect conformational changes in bulk samples of protein. Extending tmFRET to monitor the conformational state of single protein molecules allows us to detect intermediate states that would be averaged out and thus unobservable in bulk samples, as well as measure the rate of conformational state transitions at equilibrium. To achieve single-molecule tmFRET measurements using a maltose binding protein (MBP) model system, we: label MBP with a metal ion at an engineered metal-binding site and with a fluorophore using cysteine-specific chemistry; specifically anchor MBP molecules to a functionalized glass coverslip; and image single MBP molecules using total-internal-reflection-fluorescence microscopy. Finally, we use quantitative image analysis to recover fluorescence parameters that we can interpret in the context of MBP’s previously well-characterized conformational transitions. Preliminary results culminating in movies of individual fluorescent MBP molecules are presented. Overall, this novel combination of tmFRET with single-molecule fluorescence is a powerful new tool for researchers seeking to probe the conformational dynamics of proteins of biological interest.