Biology subjects

Kirkpatrick, S. L.

Publications and source records attributed to Kirkpatrick, S. L..

5 recordsLinked to original sources

bioRxiv2025

Circadian Rhythms of Whole-Body Metabolite Abundance in Drosophila are Largely Driven by Time of Feeding

Organisms exhibit daily oscillations in metabolite abundance. These oscillations could arise from circadian clock control of metabolic pathways in peripheral tissues, or secondary to rhythmic food intake, which is primarily controlled by central circadian clocks in the brain. To determine the relative contribution of central and peripheral clocks and behavioral cycles to metabolic rhythms in the fruit fly, Drosophila melanogaster, we conducted large-scale metabolite profiling with fine temporal resolution across multiple days in control flies with intact molecular clocks and in flies in which we used the CRISPR/Cas9 gene editing system to specifically eliminate molecular circadian clock function in the fat body, a peripheral metabolic tissue, or the brain. As these latter flies lack feeding rhythms due to central circadian clock dysfunction, we also included an experimental cohort of flies lacking central brain clocks but subjected to time-restricted feeding (TRF) protocols to impose feeding rhythms. Single-nuclei RNA sequencing confirmed selective molecular clock elimination following fat body manipulations, which was associated with predicted alterations in clock gene expression and an attenuation of time-of-day differences in the abundance of fat body transcripts involved in key metabolic pathways. Interestingly, we identified few rhythmically expressed metabolites in flies that were allowed ad libitum food access, and rhythms of metabolite abundance were not drastically altered by tissue-specific molecular clock disruption. In contrast, we found that flies lacking brain clocks but raised on TRF exhibited a profound increase in cyclic metabolites. These findings suggest that whole-body metabolic rhythms in Drosophila are more strongly regulated by feeding cycles than by direct circadian clock control of metabolic pathways despite the presence of metabolic genes that exhibit local-clock dependent modulation of expression across the day.

molecular biology↗

bioRxiv2024

Atp1a2 and Kcnj9 are candidate genes underlying oxycodone behavioral sensitivity and withdrawal in C57BL/6 substrains

Opioid use disorder is heritable, yet its genetic etiology is largely unknown. C57BL/6J and C57BL/6NJ mouse substrains exhibit phenotypic diversity in the context of limited genetic diversity which together can facilitate genetic discovery. Here, we found C57BL/6NJ mice were less sensitive to oxycodone (OXY)-induced locomotor activation versus C57BL/6J mice in a conditioned place preference paradigm. Narrow-sense heritability was estimated at 0.22-0.31, implicating suitability for genetic analysis. Quantitative trait locus (QTL) mapping in an F2 cross identified a chromosome 1 QTL explaining 7-12% of the variance in OXY locomotion and anxiety-like withdrawal in the elevated plus maze. A second QTL for EPM withdrawal behavior on chromosome 5 near Gabra2 (alpha-2 subunit of GABA-A receptor) explained 9% of the variance. To narrow the chromosome 1 locus, we generated recombinant lines spanning 163-181 Mb, captured the QTL for OXY locomotor traits and withdrawal, and fine-mapped a 2.45-Mb region (170.16-172.61 Mb). Transcriptome analysis identified five, localized striatal cis-eQTL transcripts and two were confirmed at the protein level (KCNJ9, ATP1A2). Kcnj9 codes for a potassium channel (GIRK3) that is a major effector of mu opioid receptor signaling. Atp1a2 codes for a subunit of a Na+/K+ ATPase enzyme that regulates neuronal excitability and shows functional adaptations following chronic opioid administration. To summarize, we identified two candidate genes underlying the physiological and behavioral properties of opioids, with direct preclinical relevance to investigators employing these widely used substrains and clinical relevance to human genetic studies of opioid use disorder.

genetics↗

bioRxiv2021

A quantitative trait variant in Gabra2 underlies increased methamphetamine stimulant sensitivity

Psychostimulant (methamphetamine, cocaine) use disorders have a genetic component that remains mostly unknown. Here, we conducted genome-wide quantitative trait locus (QTL) analysis of methamphetamine stimulant sensitivity. To facilitate gene identification, we employed a Reduced Complexity Cross between closely related C57BL/6 mouse substrains and examined maximum speed and distance traveled over 30 min following methamphetamine (2 mg/kg, i.p.). For maximum methamphetamine-induced speed following the second and third administration, we identified a single genome-wide significant QTL on chromosome 11 that peaked near the Cyfip2 locus [LOD = 3.5, 4.2; peak = 21 cM (36 Mb)]. For methamphetamine-induced distance traveled, we identified a single genome-wide significant QTL on chromosome 5 that peaked near a functional intronic indel in Gabra2 that codes for the alpha-2 subunit of the GABA-A receptor [LOD = 5.2; peak = 35 cM (67 Mb)]. Striatal cis-expression QTL mapping corroborated Gabra2 as a functional candidate gene underlying methamphetamine-induced distance traveled. CRISPR/Cas9-mediated correction of the mutant intronic deletion on the C57BL/6J background to the wild-type C57BL/6NJ allele was sufficient to reduce methamphetamine-induced locomotor activity toward the wild-type C57BL/6NJ-like level, thus validating the quantitative trait variant (QTV). These studies demonstrate the power and efficiency of Reduced Complexity Crosses in identifying causal genes and variants underlying complex traits. Functionally restoring Gabra2 expression decreased methamphetamine stimulant sensitivity and supports preclinical and human genetic studies implicating the GABA-A receptor in psychostimulant addiction-relevant traits. Importantly, our findings have major implications for investigators studying psychostimulants in the C57BL/6J strain - the gold standard strain in biomedical research.

genetics↗

bioRxiv2018

C57BL/6 substrain differences in inflammatory and neuropathic nociception and genetic mapping of a major quantitative trait locus underlying acute thermal nociception

Sensitivity to different pain modalities has a genetic basis that remains largely unknown. Employing closely related inbred mouse substrains can facilitate gene mapping of nociceptive behaviors in preclinical pain models. We previously reported enhanced sensitivity to acute thermal nociception in C57BL/6J (B6J) versus C57BL/6N (B6N) substrains. Here, we expanded on nociceptive phenotypes and observed an increase in formalin-induced inflammatory nociceptive behaviors and paw diameter in B6J versus B6N mice (Charles River Laboratories). No strain differences were observed in mechanical or thermal hypersensitivity or in edema following the Complete Freunds Adjuvant (CFA) model of inflammatory pain, indicating specificity in the inflammatory nociceptive stimulus. In the chronic nerve constriction injury (CCI), a model of neuropathic pain, no strain differences were observed in baseline mechanical threshold or in mechanical hypersensitivity up to one month post-CCI. We replicated the enhanced thermal nociception in the 52.5{degrees}C hot plate test in B6J versus B6N mice from The Jackson Laboratory. Using a B6J x B6N-F2 cross (N=164), we mapped a major quantitative trait locus (QTL) underlying hot plate sensitivity to chromosome 7 that peaked at 26 Mb (LOD=3.81, p<0.01; 8.74 Mb-36.50 Mb) that was more pronounced in males. Genes containing expression QTLs (eQTLs) associated with the peak nociceptive marker that are implicated in pain and inflammation include Ryr1, Cyp2a5, Pou2f2, Clip3, Sirt2, Actn4, and Ltbp4 (FDR < 0.05). Future studies involving positional cloning and gene editing will determine the quantitative trait gene(s) and potential pleiotropy of this locus across pain modalities.

genetics↗

bioRxiv2018

Cyfip1 haploinsufficiency increases compulsive-like behavior and paternally inherited palatable food intake: Implications for Prader-Willi Syndrome

Binge eating (BE) is a heritable trait associated with eating disorders and involves rapid consumption of large quantities of food. We identified cytoplasmic FMRP-interacting protein 2 (Cyfip2) as a major genetic factor underlying BE and concomitant compulsive-like behaviors in mice. CYFIP2 is a gene homolog of CYFIP1 - one of four paternally-deleted genes in patients with the more severe Type I Prader-Willi Syndrome (PWS). PWS is a neurodevelopmental disorder where 70% of cases involve paternal deletion of 15q11-q13. PWS symptoms include hyperphagia, obesity (if untreated), cognitive deficits, and obsessive-compulsive behaviors. We tested whether Cyfip1 haploinsufficiency (+/-) would enhance premorbid compulsive-like behavior and palatable food (PF) intake in a parent-of-origin-selective manner. We tested Cyfip1+/- mice on a C57BL/6N (N) background that were homozygous for the BE-associated missense mutation in Cyfip2 (S968F) as well as mice that we backcrossed to homozygosity for the C57BL/6J (J) allele at Cyfip2 (Cyfip2J/J). Cyfip1+/- mice showed increased compulsive-like behavior on both backgrounds, increased PF consumption on the Cyfip2N/N background in a paternally-enhanced manner, and decreased PF consumption in male Cyfip1+/- mice on the Cyfip2J/J background in a maternally selective manner. In the hypothalamus, there was a maternally-enhanced reduction of Cyfip1 transcription, but a paternally-enhanced reduction in CYFIP1 protein. In the nucleus accumbens, there was a maternally-enhanced reduction in CYFIP1 protein. Together, increased compulsive-like behavior, parent-of-origin-, and genetic background-dependent effects of Cyfip1 haploinsufficiency on PF consumption implicate CYFIP1 in behaviors in neurodevelopmental disorders involving reduced expression of CYFIP1, including PWS, Fragile X Syndrome, and 15q11.2 Microdeletion Syndrome.

genetics↗