Faculty Bibliography

A synthetic approach to several sesterterpenoids containing an isopropyl trans-hydrindane system is presented. Its most remarkable feature is the stereochemical diversification of a common precursor through the choice of different hydrogenation conditions.

Advances in synthetic chemistry, structural biology, molecular modelling and molecular cloning have enabled the systematic functional manipulation of transmembrane proteins. By combining genetically manipulated proteins with light-sensitive ligands, innately 'blind' neurobiological receptors can be converted into photoreceptors, which allows them to be photoregulated with high spatiotemporal precision. Here, we present the optochemical control of neuronal nicotinic acetylcholine receptors (nAChRs) with photoswitchable tethered agonists and antagonists. Using structure-based design, we produced heteromeric alpha3beta4 and alpha4beta2 nAChRs that can be activated or inhibited with deep-violet light, but respond normally to acetylcholine in the dark. The generation of these engineered receptors should facilitate investigation of the physiological and pathological functions of neuronal nAChRs and open a general pathway to photosensitizing pentameric ligand-gated ion channels.

BACKGROUND: Stress sensitivity and HPA axis activity may be relevant to the development and expression of psychotic disorders. Cortisol secretion has been associated with positive symptoms both in patients with psychotic disorders and in young people at clinical risk for psychosis. Herein, we aimed to replicate these findings, to determine which positive symptoms may be associated with cortisol levels, and to explore any associations with affective symptoms and impaired stress tolerance. METHODS: Thirty-one clinical high risk patients were evaluated in cross-section for associations between salivary cortisol levels upon clinic entry at 11 am, demographic variables, and clinical symptoms. RESULTS: Salivary cortisol levels were unrelated to medication exposure or demographics, except for higher levels in the ten females studied. Salivary cortisol bore no relationship to overall positive symptom severity but was associated with anxiety, as well as with suspiciousness and impaired stress tolerance, which were themselves highly intercorrelated. CONCLUSIONS: Cortisol secretion in the context of a putative novel social situation (i.e. clinic entry) may be a biological correlate of suspiciousness, impaired stress tolerance and affective symptoms in individuals vulnerable to developing psychosis. These associations are consistent with findings from experience sampling studies in individuals at risk for psychosis as well as basic studies of animal models of schizophrenia.

BACKGROUND: Clinical and epidemiological studies suggest an association between cannabis use and psychosis but this relationship remains controversial. METHOD: Clinical high-risk (CHR) subjects (age 12-22 years) with attenuated positive symptoms of psychosis (CHR+, n=101) were compared to healthy controls (HC, n=59) on rates of substance use, including cannabis. CHR+ subjects with and without lifetime cannabis use (and abuse) were compared on prodromal symptoms and social/role functioning at baseline. Participants were followed an average of 2.97 years to determine psychosis conversion status and functional outcome. RESULTS: At baseline, CHR+ subjects had significantly higher rates of lifetime cannabis use than HC. CHR+ lifetime cannabis users (n=35) were older (p=0.015, trend), more likely to be Caucasian (p=0.002), less socially anhedonic (p<0.001) and had higher Global Functioning: Social (GF:Social) scores (p<0.001) than non-users (n=61). CHR+ cannabis users continued to have higher social functioning than non-users at follow-up (p<0.001) but showed no differences in role functioning. A small sample of CHR+ cannabis abusers (n=10) showed similar results in that abusers were older (p=0.008), less socially anhedonic (p=0.017, trend) and had higher baseline GF:Social scores (p=0.006) than non-abusers. Logistic regression analyses revealed that conversion to psychosis in CHR+ subjects (n=15) was not related to lifetime cannabis use or abuse. CONCLUSIONS: The current data do not indicate that low to moderate lifetime cannabis use is a major contributor to psychosis or poor social and role functioning in clinical high-risk youth with attenuated positive symptoms of psychosis.

Metabolic syndrome is a condition that typically includes central obesity, insulin resistance, glucose intolerance, dyslipidemia, and hypertension. Disruption of the hypothalamic-pituitary-adrenal axis, a regulator of corticosterone secretion, occurs in some cases of metabolic syndrome and obesity, and Cushing hypercortisolemia is associated with obesity and metabolic disorders. We therefore assessed anatomic and clinical pathology in C57BL/6NCrl mice to evaluate the effects of chronic corticosterone in the drinking water at doses of 25, 50, and 100 mug/mL for 25 d. Treated mice developed obesity, glucose intolerance, electrolyte aberrations, and dyslipidemia that were dose-dependent and most severe in the 100-mu;g/mL treatment group. To evaluate return to normal function, additional C57BL/6NCrl mice received corticosterone-free water for 2 wk after the 25-d treatment period. According to results of gross examination, mice appeared to recover within days of exogenous corticosterone withdrawal; however, adrenal gland vacuolation and protein, lipid, and electrolyte abnormalities persisted. Together, these findings support chronic corticosterone exposure through the drinking water as a potentially useful, noninvasive method to induce some features of metabolic syndrome.

Changes in neural activity influence synaptic plasticity/scaling, gene expression, and epigenetic modifications. We present the first evidence that short-term and persistent changes in neural activity can alter adenosine-to-inosine (A-to-I) RNA editing, a post-transcriptional site-specific modification found in several neuron-specific transcripts. In rat cortical neuron cultures, activity-dependent changes in A-to-I RNA editing in coding exons are present after 6 hr of high potassium depolarization but not after 1 hr and require calcium entry into neurons. When treatments are extended from hours to days, we observe a negative feedback phenomenon: Chronic depolarization increases editing at many sites and chronic silencing decreases editing. We present several different modulations of neural activity that change the expression of different mRNA isoforms through editing.

Transcription activator-like effectors (TALEs) are a class of naturally occurring DNA-binding proteins found in the plant pathogen Xanthomonas sp. The DNA-binding domain of each TALE consists of tandem 34-amino acid repeat modules that can be rearranged according to a simple cipher to target new DNA sequences. Customized TALEs can be used for a wide variety of genome engineering applications, including transcriptional modulation and genome editing. Here we describe a toolbox for rapid construction of custom TALE transcription factors (TALE-TFs) and nucleases (TALENs) using a hierarchical ligation procedure. This toolbox facilitates affordable and rapid construction of custom TALE-TFs and TALENs within 1 week and can be easily scaled up to construct TALEs for multiple targets in parallel. We also provide details for testing the activity in mammalian cells of custom TALE-TFs and TALENs using quantitative reverse-transcription PCR and Surveyor nuclease, respectively. The TALE toolbox described here will enable a broad range of biological applications.