The regulatory landscape of multiple brain regions in outbred heterogeneous stock rats.

Heterogeneous Stock (HS) rats are a genetically diverse outbred rat population that is widely used for studying genetics of behavioral and physiological traits. Mapping Quantitative Trait Loci (QTL) associated with transcriptional changes would help to identify mechanisms underlying these traits. We generated genotype and transcriptome data for five brain regions from 88 HS rats. We identified 21 392 cis-QTLs associated with expression and splicing changes across all five brain regions and validated their effects using allele specific expression data. We identified 80 cases where eQTLs were colocalized with genome-wide association study (GWAS) results from nine physiological traits. Comparing our dataset to human data from the Genotype-Tissue Expression (GTEx) project, we found that the HS rat data yields twice as many significant eQTLs as a similarly sized human dataset. We also identified a modest but highly significant correlation between genetic regulatory variation among orthologous genes. Surprisingly, we found less genetic variation in gene regulation in HS rats relative to humans, though we still found eQTLs for the orthologs of many human genes for which eQTLs had not been found. These data are available from the RatGTEx data portal (RatGTEx.org) and will enable new discoveries of the genetic influences of complex traits.

Genome-wide association mapping of ethanol sensitivity in the Diversity Outbred mouse population.

BACKGROUND: A strong predictor for the development of alcohol use disorder (AUD) is altered sensitivity to the intoxicating effects of alcohol. Individual differences in the initial sensitivity to alcohol are controlled in part by genetic factors. Mice offer a powerful tool to elucidate the genetic basis of behavioral and physiological traits relevant to AUD, but conventional experimental crosses have only been able to identify large chromosomal regions rather than specific genes. Genetically diverse, highly recombinant mouse populations make it possible to observe a wider range of phenotypic variation, offer greater mapping precision, and thus increase the potential for efficient gene identification. METHODS: We have taken advantage of the Diversity Outbred (DO) mouse population to identify and precisely map quantitative trait loci (QTL) associated with ethanol sensitivity. We phenotyped 798 male J:DO mice for three measures of ethanol sensitivity: ataxia, hypothermia, and loss of the righting response. We used high-density MegaMUGA and GigaMUGA to obtain genotypes ranging from 77,808 to 143,259 SNPs. We also performed RNA sequencing in striatum to map expression QTLs and identify gene expression-trait correlations. We then applied a systems genetic strategy to identify narrow QTLs and construct the network of correlations that exists between DNA sequence, gene expression values, and ethanol-related phenotypes to prioritize our list of positional candidate genes. RESULTS: We observed large amounts of phenotypic variation with the DO population and identified suggestive and significant QTLs associated with ethanol sensitivity on chromosomes 1, 2, and 16. The implicated regions were narrow (4.5-6.9 Mb in size) and each QTL explained ~4-5% of the variance. CONCLUSIONS: Our results can be used to identify alleles that contribute to AUD in humans, elucidate causative biological mechanisms, or assist in the development of novel therapeutic interventions.

Optogenetic regulation of transcription.

Optogenetics has become widely recognized for its success in real-time control of brain neurons by utilizing non-mammalian photosensitive proteins to open or close membrane channels. Here we review a less well known type of optogenetic constructs that employs photosensitive proteins to transduce the signal to regulate gene transcription, and its possible use in medicine. One of the problems with existing gene therapies is that they could remain active indefinitely while not allowing regulated transgene production on demand. Optogenetic regulation of transcription (ORT) could potentially be used to regulate the production of a biological drug in situ, by repeatedly applying light to the tissue, and inducing expression of therapeutic transgenes when needed. Red and near infrared wavelengths, which are capable of penetration into tissues, have potential for therapeutic applications. Existing ORT systems are reviewed herein with these considerations in mind.

Chronic central nervous system expression of HIV-1 Tat leads to accelerated rarefaction of neocortical capillaries and loss of red blood cell velocity heterogeneity.

OBJECTIVES: HIV-1 infection of the CNS is associated with impairment of CBF and neurocognitive function, and accelerated signs of aging. As normal aging is associated with rarefaction of the cerebral vasculature, we set out to examine chronic viral effects on the cerebral vasculature. METHODS: DOX-inducible HIV-1 Tat-tg and WT control mice were used. Animals were treated with DOX for three weeks or five to seven months. Cerebral vessel density and capillary segment length were determined from quantitative image analyses of sectioned cortical tissue. In addition, movement of red blood cells in individual capillaries was imaged in vivo using multiphoton microscopy, to determine RBCV and flux. RESULTS: Mean RBCV was not different between Tat-tg mice and age-matched WT controls. However, cortical capillaries from Tat-tg mice showed a significant loss of RBCV heterogeneity and increased RBCF that was attributed to a marked decrease in total cortical capillary length (35-40%) compared to WT mice. CONCLUSIONS: Cerebrovascular rarefaction is accelerated in HIV-1 Tat-transgenic mice, and this is associated with alterations in red cell blood velocity. These changes may have relevance to the pathogenesis of HIV-associated neurocognitive disorders in an aging HIV-positive population.

Fatigability disrupts cognitive processes' regulation of inflammatory reactivity in old age.

OBJECTIVE: High fatigability, a dysfunctional adaption to fatigue, may lead to difficulties performing otherwise regularly encountered cognitive activities and may be related to pro-inflammatory reactivity. The purpose of the study was to investigate the effect of fatigability on cognitive processes and inflammatory response after an acute cognitive stress task in older adults. METHODS: In an observational stress reactivity study conducted in a light- and temperature-controlled laboratory, we measured IL-6, self-reported acute fatigue, and frontally oriented cognitive processes in 55 community-dwelling individuals aged 75 years or older as part of a demanding set of cognitive tasks intended to induce stress. RESULTS: Subjects were classified into groups of low and high fatigability based on cluster analysis of their self-report acute fatigue before and after the cognitive tasks. The two clusters were comparable on levels of baseline IL-6 and cognitive processes; however, the high fatigability cluster had significantly higher levels of IL-6 response than the low fatigability cluster. After controlling for multiple covariates, fatigability moderated the relationship between speed of processing and IL-6 reactivity. Further exploratory analyses indicated significant adverse associations between speed of processing and attention and IL-6 reactivity in the group with low but not high fatigability. CONCLUSION: Although observational, these data are consistent with the notion that pro-inflammatory states in older adults might be reduced by improvements in cognitive processes. Because fatigability was associated with increased acute inflammatory response and disrupted the normal stress regulation provided by the cognitive processes, future randomized studies might examine whether fatigability alleviation reduces IL-6.

Genome-wide association study for age-related hearing loss in CFW mice.

Age-related hearing impairment is the most common cause of hearing loss and is one of the most prevalent conditions affecting the elderly globally. It is influenced by a combination of environmental and genetic factors. The mouse and human inner ears are functionally and genetically homologous. Investigating the genetic basis of age-related hearing loss (ARHL) in an outbred mouse model may lead to a better understanding of the molecular mechanisms of this condition. We used Carworth Farms White (CFW) outbred mice, because they are genetically diverse and exhibit variation in the onset and severity of ARHL. The goal of this study was to identify genetic loci involved in regulating ARHL. Hearing at a range of frequencies was measured using Auditory Brainstem Response (ABR) thresholds in 946 male and female CFW mice at the age of 1, 6, and 10 months. We obtained genotypes at 4.18 million single nucleotide polymorphisms (SNP) using low-coverage (mean coverage 0.27x) whole-genome sequencing followed by imputation using STITCH. To determine the accuracy of the genotypes we sequenced 8 samples at >30x coverage and used calls from those samples to estimate the discordance rate, which was 0.45%. We performed genetic analysis for the ABR thresholds for each frequency at each age, and for the time of onset of deafness for each frequency. The SNP heritability ranged from 0 to 42% for different traits. Genome-wide association analysis identified several regions associated with ARHL that contained potential candidate genes, including Dnah11, Rapgef5, Cpne4, Prkag2, and Nek11. We confirmed, using functional study, that Prkag2 deficiency causes age-related hearing loss at high frequency in mice; this makes Prkag2 a candidate gene for further studies. This work helps to identify genetic risk factors for ARHL and to define novel therapeutic targets for the treatment and prevention of ARHL.

Environmental enrichment promotes adaptive responding during tests of behavioral regulation in male heterogeneous stock rats.

Organisms must regulate their behavior flexibly in the face of environmental challenges. Failure can lead to a host of maladaptive behavioral traits associated with a range of neuropsychiatric disorders, including attention deficit hyperactivity disorder, autism, and substance use disorders. This maladaptive dysregulation of behavior is influenced by genetic and environmental factors. For example, environmental enrichment produces beneficial neurobehavioral effects in animal models of such disorders. The present study determined the effects of environmental enrichment on a range of measures related to behavioral regulation using a large cohort of male, outbred heterogeneous stock (HS) rats as subjects. Subjects were reared from late adolescence onwards either in pairs in standard housing with minimal enrichment (n = 200) or in groups of 16 in a highly enriched environment consisting of a large multi-level cage filled with toys, running wheels, and shelters (n = 64). Rats were subjected to a battery of tests, including: (i) locomotor response to novelty, (ii) light reinforcement, (iii) social reinforcement, (iv) reaction time, (v) a patch-depletion foraging test, (vi) Pavlovian conditioned approach, (vii) conditioned reinforcement, and (viii) cocaine conditioned cue preference. Results indicated that rats housed in the enriched environment were able to filter out irrelevant stimuli more effectively and thereby regulate their behavior more efficiently than standard-housing rats. The dramatic impact of environmental enrichment suggests that behavioral studies using standard housing conditions may not generalize to more complex environments that may be more ethologically relevant.

Genomic Loci Influencing Cue-Reactivity in Heterogeneous Stock Rats.

Addiction vulnerability is associated with the tendency to attribute incentive salience to reward predictive cues; both addiction and the attribution of incentive salience are influenced by environmental and genetic factors. To characterize the genetic contributions to incentive salience attribution, we performed a genome-wide association study (GWAS) in a cohort of 1,645 genetically diverse heterogeneous stock (HS) rats. We tested HS rats in a Pavlovian conditioned approach task, in which we characterized the individual responses to food-associated stimuli ("cues"). Rats exhibited either cue-directed "sign-tracking" behavior or food-cup directed "goal-tracking" behavior. We then used the conditioned reinforcement procedure to determine whether rats would perform a novel operant response for unrewarded presentations of the cue. We found that these measures were moderately heritable (SNP heritability, h2 = .189-.215). GWAS identified 14 quantitative trait loci (QTLs) for 11 of the 12 traits we examined. Interval sizes of these QTLs varied widely. 7 traits shared a QTL on chromosome 1 that contained a few genes (e.g. Tenm4, Mir708) that have been associated with substance use disorders and other mental health traits in humans. Other candidate genes (e.g. Wnt11, Pak1) in this region had coding variants and expression-QTLs in mesocorticolimbic regions of the brain. We also conducted a Phenome-Wide Association Study (PheWAS) on other behavioral measures in HS rats and found that regions containing QTLs on chromosome 1 were also associated with nicotine self-administration in a separate cohort of HS rats. These results provide a starting point for the molecular genetic dissection of incentive salience and provide further support for a relationship between attribution of incentive salience and drug abuse-related traits.

Genome-wide association study reveals multiple loci for nociception and opioid consumption behaviors associated with heroin vulnerability in outbred rats.

The increased prevalence of opioid use disorder (OUD) makes it imperative to disentangle the biological mechanisms contributing to individual differences in OUD vulnerability. OUD shows strong heritability, however genetic variants contributing toward vulnerability remain poorly defined. We performed a genome-wide association study using over 850 male and female heterogeneous stock (HS) rats to identify genes underlying behaviors associated with OUD such as nociception, as well as heroin-taking, extinction and seeking behaviors. By using an animal model of OUD, we were able to identify genetic variants associated with distinct OUD behaviors while maintaining a uniform environment, an experimental design not easily achieved in humans. Furthermore, we used a novel non-linear network-based clustering approach to characterize rats based on OUD vulnerability to assess genetic variants associated with OUD susceptibility. Our findings confirm the heritability of several OUD-like behaviors, including OUD susceptibility. Additionally, several genetic variants associated with nociceptive threshold prior to heroin experience, heroin consumption, escalation of intake, and motivation to obtain heroin were identified. Tom1 , a microglial component, was implicated for nociception. Several genes involved in dopaminergic signaling, neuroplasticity and substance use disorders, including Brwd1 , Pcp4, Phb1l2 and Mmp15 were implicated for the heroin traits. Additionally, an OUD vulnerable phenotype was associated with genetic variants for consumption and break point, suggesting a specific genetic contribution for OUD-like traits contributing to vulnerability. Together, these findings identify novel genetic markers related to the susceptibility to OUD-relevant behaviors in HS rats.

A revamped rat reference genome improves the discovery of genetic diversity in laboratory rats.

The seventh iteration of the reference genome assembly for Rattus norvegicus-mRatBN7.2-corrects numerous misplaced segments and reduces base-level errors by approximately 9-fold and increases contiguity by 290-fold compared with its predecessor. Gene annotations are now more complete, improving the mapping precision of genomic, transcriptomic, and proteomics datasets. We jointly analyzed 163 short-read whole-genome sequencing datasets representing 120 laboratory rat strains and substrains using mRatBN7.2. We defined ∼20.0 million sequence variations, of which 18,700 are predicted to potentially impact the function of 6,677 genes. We also generated a new rat genetic map from 1,893 heterogeneous stock rats and annotated transcription start sites and alternative polyadenylation sites. The mRatBN7.2 assembly, along with the extensive analysis of genomic variations among rat strains, enhances our understanding of the rat genome, providing researchers with an expanded resource for studies involving rats.

Y and Mitochondrial Chromosomes in the Heterogeneous Stock Rat Population.

Genome-wide association studies typically evaluate the autosomes and sometimes the X Chromosome, but seldom consider the Y or mitochondrial Chromosomes. We genotyped the Y and mitochondrial chromosomes in heterogeneous stock rats (Rattus norvegicus), which were created in 1984 by intercrossing eight inbred strains and have subsequently been maintained as an outbred population for 100 generations. As the Y and mitochondrial Chromosomes do not recombine, we determined which founder had contributed these chromosomes for each rat, and then performed association analysis for all complex traits (n=12,055; intersection of 12,116 phenotyped and 15,042 haplotyped rats). We found the eight founders had 8 distinct Y and 4 distinct mitochondrial Chromosomes, however only two of each were observed in our modern heterogeneous stock rat population (Generations 81-97). Despite the unusually large sample size, the p-value distribution did not deviate from expectations; there were no significant associations for behavioral, physiological, metabolome, or microbiome traits after correcting for multiple comparisons. However, both Y and mitochondrial Chromosomes were strongly associated with expression of a few genes located on those chromosomes, which provided a positive control. Our results suggest that within modern heterogeneous stock rats there are no Y and mitochondrial Chromosomes differences that strongly influence behavioral or physiological traits. These results do not address other ancestral Y and mitochondrial Chromosomes that do not appear in modern heterogeneous stock rats, nor do they address effects that may exist in other rat populations, or in other species.

Causal Genetic Loci for a Motivated Behavior Spectrum Harbor Psychiatric Risk Genes.

Behavioral diversity is critical for population fitness. Individual differences in risk-taking are observed across species, but underlying genetic mechanisms and conservation are largely unknown. We examined dark avoidance in larval zebrafish, a motivated behavior reflecting an approach-avoidance conflict. Brain-wide calcium imaging revealed significant neural activity differences between approach-inclined versus avoidance-inclined individuals. We used a population of ∼6,000 to perform the first genome-wide association study (GWAS) in zebrafish, which identified 34 genomic regions harboring many genes that are involved in synaptic transmission and human psychiatric diseases. We used CRISPR to study several causal genes: serotonin receptor-1b ( htr1b ), nitric oxide synthase-1 ( nos1 ), and stress-induced phosphoprotein-1 ( stip1 ). We further identified 52 conserved elements containing 66 GWAS significant variants. One encoded an exonic regulatory element that influenced tissue-specific nos1 expression. Together, these findings reveal new genetic loci and establish a powerful, scalable animal system to probe mechanisms underlying motivation, a critical dimension of psychiatric diseases.

Automated quantitative trait locus analysis (AutoQTL).

Background: Quantitative Trait Locus (QTL) analysis and Genome-Wide Association Studies (GWAS) have the power to identify variants that capture significant levels of phenotypic variance in complex traits. However, effort and time are required to select the best methods and optimize parameters and pre-processing steps. Although machine learning approaches have been shown to greatly assist in optimization and data processing, applying them to QTL analysis and GWAS is challenging due to the complexity of large, heterogenous datasets. Here, we describe proof-of-concept for an automated machine learning approach, AutoQTL, with the ability to automate many complex decisions related to analysis of complex traits and generate diverse solutions to describe relationships that exist in genetic data. Results: Using a dataset of 18 putative QTL from a large-scale GWAS of body mass index in the laboratory rat, Rattus norvegicus , AutoQTL captures the phenotypic variance explained under a standard additive model while also providing evidence of non-additive effects including deviations from additivity and 2-way epistatic interactions from simulated data via multiple optimal solutions. Additionally, feature importance metrics provide different insights into the inheritance models and predictive power of multiple GWAS-derived putative QTL. Conclusions: This proof-of-concept illustrates that automated machine learning techniques can be applied to genetic data and has the potential to detect both additive and non-additive effects via various optimal solutions and feature importance metrics. In the future, we aim to expand AutoQTL to accommodate omics-level datasets with intelligent feature selection strategies.

Environmental enrichment promotes adaptive responding during tests of behavioral regulation in male heterogeneous stock rats.

Organisms must regulate their behavior flexibly in the face of environmental challenges. Failure can lead to a host of maladaptive behavioral traits associated with a range of neuropsychiatric disorders, including attention deficit hyperactivity disorder, autism, and substance use disorders. This maladaptive dysregulation of behavior is influenced by genetic and environmental factors. For example, environmental enrichment produces beneficial neurobehavioral effects in animal models of such disorders. The present study determined the effects of environmental enrichment on a range of measures related to behavioral regulation using a large cohort of male, outbred heterogeneous stock (HS) rats as subjects to mimic the genetic variability found in the human population. Subjects were reared from late adolescence onwards either in pairs in standard housing with minimal enrichment (n=200) or in groups of 16 in a highly enriched environment consisting of a large multi-level cage filled with toys, running wheels, and shelters (n=64). Rats were subjected to a battery of tests, including: (i) locomotor response to novelty, (iI) light reinforcement, (iii) social reinforcement, (iv) reaction time, (v) a patch-depletion foraging test, (vi) Pavlovian conditioned approach, (vii) conditioned reinforcement, and (viii) cocaine conditioned cue preference. Results indicated that rats housed in the enriched environment were able to filter out irrelevant stimuli more effectively and thereby regulate their behavior more efficiently than standard-housing rats. The dramatic impact of environmental enrichment suggests that behavioral studies using standard housing conditions may not generalize to more complex environments that may be more ethologically relevant.

Genome-wide association study of delay discounting in Heterogenous Stock rats.

Delay discounting refers to the behavioral tendency to devalue rewards as a function of their delay in receipt. Heightened delay discounting has been associated with substance use disorders, as well as multiple co-occurring psychopathologies. Genetic studies in humans and animal models have established that delay discounting is a heritable trait, but only a few specific genes have been associated with delay discounting. Here, we aimed to identify novel genetic loci associated with delay discounting through a genome-wide association study (GWAS) using Heterogenous Stock rats, a genetically diverse outbred population derived from eight inbred founder strains. We assessed delay discounting in 650 male and female rats using an adjusting amount procedure in which rats chose between smaller immediate sucrose rewards or a larger reward at variable delays. Preference switch points were calculated for each rat and both exponential and hyperbolic functions were fitted to these indifference points. Area under the curve (AUC) and the discounting parameter k of both functions were used as delay discounting measures. GWAS for AUC, exponential k, and indifference points for a short delay identified significant loci on chromosomes 20 and 14. The gene Slc35f1, which encodes a member of the solute carrier family of nucleoside sugar transporters, was the only gene within the chromosome 20 locus. That locus also contained an eQTL for Slc35f1, suggesting that heritable differences in the expression of that gene might be responsible for the association with behavior. The gene Adgrl3, which encodes a member of the latrophilin family of G-protein coupled receptors, was the only gene within the chromosome 14 locus. These findings implicate novel genes in delay discounting and highlight the need for further exploration.

Automated quantitative trait locus analysis (AutoQTL).

BACKGROUND: Quantitative Trait Locus (QTL) analysis and Genome-Wide Association Studies (GWAS) have the power to identify variants that capture significant levels of phenotypic variance in complex traits. However, effort and time are required to select the best methods and optimize parameters and pre-processing steps. Although machine learning approaches have been shown to greatly assist in optimization and data processing, applying them to QTL analysis and GWAS is challenging due to the complexity of large, heterogenous datasets. Here, we describe proof-of-concept for an automated machine learning approach, AutoQTL, with the ability to automate many complicated decisions related to analysis of complex traits and generate solutions to describe relationships that exist in genetic data. RESULTS: Using a publicly available dataset of 18 putative QTL from a large-scale GWAS of body mass index in the laboratory rat, Rattus norvegicus, AutoQTL captures the phenotypic variance explained under a standard additive model. AutoQTL also detects evidence of non-additive effects including deviations from additivity and 2-way epistatic interactions in simulated data via multiple optimal solutions. Additionally, feature importance metrics provide different insights into the inheritance models and predictive power of multiple GWAS-derived putative QTL. CONCLUSIONS: This proof-of-concept illustrates that automated machine learning techniques can complement standard approaches and have the potential to detect both additive and non-additive effects via various optimal solutions and feature importance metrics. In the future, we aim to expand AutoQTL to accommodate omics-level datasets with intelligent feature selection and feature engineering strategies.

Genome-wide association studies of human and rat BMI converge on synapse, epigenome, and hormone signaling networks.

A vexing observation in genome-wide association studies (GWASs) is that parallel analyses in different species may not identify orthologous genes. Here, we demonstrate that cross-species translation of GWASs can be greatly improved by an analysis of co-localization within molecular networks. Using body mass index (BMI) as an example, we show that the genes associated with BMI in humans lack significant agreement with those identified in rats. However, the networks interconnecting these genes show substantial overlap, highlighting common mechanisms including synaptic signaling, epigenetic modification, and hormonal regulation. Genetic perturbations within these networks cause abnormal BMI phenotypes in mice, too, supporting their broad conservation across mammals. Other mechanisms appear species specific, including carbohydrate biosynthesis (humans) and glycerolipid metabolism (rodents). Finally, network co-localization also identifies cross-species convergence for height/body length. This study advances a general paradigm for determining whether and how phenotypes measured in model species recapitulate human biology.

An exponential increase in QTL detection with an increased sample size.

Power analyses are often used to determine the number of animals required for a genome-wide association study (GWAS). These analyses are typically intended to estimate the sample size needed for at least 1 locus to exceed a genome-wide significance threshold. A related question that is less commonly considered is the number of significant loci that will be discovered with a given sample size. We used simulations based on a real data set that consisted of 3,173 male and female adult N/NIH heterogeneous stock rats to explore the relationship between sample size and the number of significant loci discovered. Our simulations examined the number of loci identified in subsamples of the full data set. The subsampling analysis was conducted for 4 traits with low (0.15 ± 0.03), medium (0.31 ± 0.03 and 0.36 ± 0.03), and high (0.46 ± 0.03) SNP-based heritabilities. For each trait, we subsampled the data 100 times at different sample sizes (500, 1,000, 1,500, 2,000, and 2,500). We observed an exponential increase in the number of significant loci with larger sample sizes. Our results are consistent with similar observations in human GWAS and imply that future rodent GWAS should use sample sizes that are significantly larger than those needed to obtain a single significant result.

A revamped rat reference genome improves the discovery of genetic diversity in laboratory rats.

The seventh iteration of the reference genome assembly for Rattus norvegicus-mRatBN7.2-corrects numerous misplaced segments and reduces base-level errors by approximately 9-fold and increases contiguity by 290-fold compared to its predecessor. Gene annotations are now more complete, significantly improving the mapping precision of genomic, transcriptomic, and proteomics data sets. We jointly analyzed 163 short-read whole genome sequencing datasets representing 120 laboratory rat strains and substrains using mRatBN7.2. We defined ~20.0 million sequence variations, of which 18.7 thousand are predicted to potentially impact the function of 6,677 genes. We also generated a new rat genetic map from 1,893 heterogeneous stock rats and annotated transcription start sites and alternative polyadenylation sites. The mRatBN7.2 assembly, along with the extensive analysis of genomic variations among rat strains, enhances our understanding of the rat genome, providing researchers with an expanded resource for studies involving rats.

Transient hypercapnia reveals an underlying cerebrovascular pathology in a murine model for HIV-1 associated neuroinflammation: role of NO-cGMP signaling and normalization by inhibition of cyclic nucleotide phosphodiesterase-5.

BACKGROUND: Cerebral blood flow (CBF) is known to be dysregulated in persons with human immunodeficiency virus 1 (HIV-1), for uncertain reasons. This is an important issue because impaired vasoreactivity has been associated with increased risk of ischemic stroke, elevated overall cardiovascular risk and cognitive impairment. METHODS: To test whether dysregulation of CBF might be due to virally-induced neuroinflammation, we used a well-defined animal model (GFAP-driven, doxycycline-inducible HIV-1 Tat transgenic (Tat-tg) mice). We then exposed the mice to a brief hypercapnic stimulus, and assessed cerebrovascular reactivity by measuring 1) changes in cerebral blood flow, using laser Doppler flowmetry and 2) changes in vascular dilation, using in vivo two-photon imaging. RESULTS: Exposure to brief hypercapnia revealed an underlying cerebrovascular pathology in Tat-tg mice. In control animals, brief hypercapnia induced a brisk increase in cortical flow (20.8% above baseline) and vascular dilation, as measured by laser Doppler flowmetry and in vivo two-photon microscopy. These responses were significantly attenuated in Tat-tg mice (11.6% above baseline), but cortical microvascular morphology and capillary density were unaltered, suggesting that the functional pathology was not secondary to vascular remodeling. To examine the mechanistic basis for the diminished cerebrovascular response to brief hypercapnia, Tat-tg mice were treated with 1) gisadenafil, a phosphodiesterase 5 (PDE5) inhibitor and 2) tetrahydrobiopterin (BH4). Gisadenafil largely restored the normal increase in cortical flow following hypercapnia in Tat-tg mice (17.5% above baseline), whereas BH4 had little effect. Gisadenafil also restored the dilation of small (<25 µm) arterioles following hypercapnia (19.1% versus 20.6% diameter increase in control and Tat-tg plus gisadenafil, respectively), although it failed to restore full dilation of larger (>25 µm) vessels. CONCLUSIONS: Taken together, these data show that HIV-associated neuroinflammation can cause cerebrovascular pathology through effects on cyclic guanosine monophosphate (cGMP) metabolism and possibly on PDE5 metabolism.