Transgenic Rat Model Research Articles

Apigenin: Exploring its neuroprotective potential in neurodegenerative disorders: Mechanisms and promising therapeutic applications

Neurodegenerative diseases pose significant challenges to global health-care systems, highlighting the urgent need for effective therapeutic strategies. Apigenin (API), a natural compound derived from various plant sources, has emerged as a potential treatment option due to its neuroprotective properties. This review article provides a comprehensive overview of API, focusing on its definition, natural sources, pharmacokinetics, and bioavailability, mechanisms of action, safety, and tolerability profile. API protects neurons in many ways, including its antioxidant activity, anti-inflammatory properties, ability to change cellular signaling pathways, ability to stop proteins from misfolding and sticking together, and ability to improve mitochondrial function. In vitro studies have demonstrated that API attenuates oxidative stress, inhibits the formation of protein aggregates, and suppresses neuroinflammation. Animal models, such as transgenic mice, rat models, and non-human primates, have provided valuable insights into the potential therapeutic benefits of API, including improved cognitive function, mitigation of motor impairments, and preservation of neuronal integrity. API’s safety and tolerability profile appears favorable based on preclinical and clinical studies, with minimal reported adverse effects. However, further investigation is required to determine optimal dosing regimens and assess potential drug interactions. In addition, while current treatment options for neurodegenerative diseases primarily focus on symptom management, API holds promise as a disease-modifying agent.

The TgF344-AD rat: behavioral and proteomic changes associated with aging and protein expression in a transgenic rat model of Alzheimer's disease

Animal models of Alzheimer's Disease (AD) are attractive tools for preclinical, prodromal drug testing. The TgF344-AD (Tg) rat exhibits cognitive deficits and 5 major hallmarks of AD. Here we show that spatial water maze (WMZ) memory deficits and proteomic differences in dorsal CA1 were present in young Tg rats. Aged learning-unimpaired (AU) and aged learning-impaired (AI) proteome associated changes were identified and differed by sex. Levels of phosphorylated tau, reactive astrocytes and microglia were significantly increased in aged Tg rats and correlated with the WMZ learning index (LI); in contrast, no significant correlation was present between amyloid plaques or insoluble Aβ levels and LI. Neuroinflammatory markers were also significantly correlated with LI and increased in female Tg rats. The anti-inflammatory marker, triggering receptor expressed on myeloid cells-2 (TREM2), was significantly reduced in aged impaired Tg rats and correlated with LI. Identifying and understanding mechanisms that allow for healthy aging by overcoming genetic drivers for AD, and/or promoting drivers for successful aging, are important for developing successful therapeutics against AD.

Effect of Heme Oxygenase-1 Depletion on Complement Regulatory Proteins Expression in the Rat.

Heme oxygenase has been implicated in the regulation of various immune responses including complement activation. Using a transgenic rat model of HO-1 depletion, the present study assessed the effect of HO-1 absence on the expression of complement regulatory proteins: decay accelerating factor (DAF), CR1-related gene/protein Y (Crry) and CD59, which act to attenuate complement activation. Protein expression was assessed by immunohistochemistry in kidney, liver, lung and spleen tissues. DAF protein was reduced in all tissues retrieved from rats lacking HO-1 (Hmox1-/-) apart from spleen tissue sections. Crry protein was also reduced, but only in Hmox1-/- kidney and liver tissue. C3b staining was augmented in the kidney and spleen from Hmox1-/- rats, suggesting that the decrease of DAF and Crry was sufficient to increase C3b deposition. The observations support an important role of HO-1 as a regulator of the complement system.

Heterozygote Dopamine Transporter Knockout Rats Display Enhanced Cocaine Locomotion in Adolescent Females

Cocaine is a powerful psychostimulant that is one of the most widely used illicit addictive. The dopamine transporter (DAT) plays a major role in mediating cocaine's reward effect. Decreases in DAT expression increase rates of drug abuse and vulnerability to comorbid psychiatric disorders. We used the novel DAT transgenic rat model to study the effects of cocaine on locomotor behaviors in adolescent rats, with an emphasis on sex. Female rats showed higher response rates to cocaine at lower acute and chronic doses, highlighting a higher vulnerability and perceived gender effects. In contrast, locomotor responses to an acute high dose of cocaine were more marked and sustained in male DAT heterozygous (HET) adolescents. The results demonstrate the augmented effects of chronic cocaine in HET DAT adolescent female rats. Knockout (KO) DAT led to a level of hyperdopaminergia which caused a marked basal hyperactivity that was unchanged, consistent with a possible ceiling effect. We suggest a role of alpha synuclein (α-syn) and PICK 1 protein expressions to the increased vulnerability in female rats. These proteins showed a lower expression in female HET and KO rats. This study highlights gender differences associated with mutations which affect DAT expression and can increase susceptibility to cocaine abuse in adolescence.

Insulin-like growth factor II receptor alpha overexpression in heart aggravates hyperglycemia-induced cardiac inflammation and myocardial necrosis.

Diabetes-induced cardiovascular complications are mainly associated with high morbidity and mortality in patients with diabetes. Insulin-like growth factor II receptor α (IGF-IIRα) is a cardiac risk factor. In this study, we hypothesized IGF-IIRα could also deteriorate diabetic heart injury. The results presented that both in vivo transgenic Sprague-Dawley rat model with specific IGF-IIRα overexpression in the heart and in vitro myocardium H9c2 cells were used to investigate the negative function of IGF-IIRα in diabetic hearts. The results showed that IGF-IIRα overexpression aided hyperglycemia in creating more myocardial injury. Pro-inflammatory factors, such as Tumor necrosis factor-alpha, Interleukin-6, Cyclooxygenase-2, Inducible nitric oxide synthase, and Nuclear factor-kappaB inflammatory cascade, are enhanced in the diabetic myocardium with cardiac-specific IGF-IIRα overexpression. Correspondingly, IGF-IIRα overexpression in the diabetic myocardium also reduced the PI3K-AKT survival axis and activated mitochondrial-dependent apoptosis. Finally, both ejection fraction and fractional shortening were be significantly decrease in diabetic rats with cardiac-specific IGF-IIRα overexpression. Overall, all results provid clear evidence that IGF-IIRα can enhance cardiac damage and is a harmful factor to the heart under high-blood glucose conditions. However, the pathophysiology of IGF-IIRα under different stresses and its downstream regulation in the heart still require further research.

Traumatic Brain Injury Exacerbates Retinal Alzheimer’s Disease Histopathology in the TgF344 Rat Model

AbstractBackgroundAlzheimer’s disease (AD) is a neurodegenerative condition of the central nervous system that affects both the brain and the retina, and can lead to visual impairment. However, the factors that modulate AD pathophysiology leading to the wide range of clinical presentations are not well understood. Notably, traumatic brain injury (TBI), which can arise from sport concussions, military combat and other causes, is associated with a 2.3‐fold higher risk of developing AD and AD‐related dementias. Yet the pathophysiological link between TBI and AD remains poorly understood. In this study we set out to evaluate the effects of TBI, AD, and their combination, on retinal histopathology, by taking advantage of a transgenic rat model (Tg‐F344‐AD) shown to recapitulate the main features of human AD pathology, and combining it with a controlled cortical impact paradigm of repetitive mild TBI (rmTBI).MethodsMale Tg‐F344‐AD rats and F344 wild‐type controls were aged to 12 months and randomly selected to undergo a two‐time unilateral controlled cortical impact (2xCCI; one week apart) that mimics rmTBI, or a sham procedure. The animals were sacrificed at four months post‐CCI to evaluate long‐term effects of the procedure, and the eyes were collected and analyzed using immunofluorescence and standard histological staining techniques to evaluate Alzheimer’s histopathology.ResultsHistopathological analyses at four months post‐impact confirm the presence of AD markers in Tg‐F344‐AD retinas. Moreover, we found that AD increases Aβ deposition and neuroinflammation in the retina, and that TBI enhances the severity of this phenotype by increasing the formation of dense core amyloid plaques.ConclusionsWe have developed and characterized a model to study the pathophysiology of AD in relation to rmTBI in the retina, which could lead to better treatments and to the development of retinal biomarkers for improved AD diagnosis.

Cerebrovascular accumulation of amyloid‐forming amylin secreted from the pancreas induces brain hypoxia

AbstractBackgroundIslet amyloid polypeptide (amylin) is a β‐cell hormone co‐secreted with insulin. Histological analyses of human brains identified amylin deposits co‐localized with parenchymal and vascular β‐amyloid in humans with vascular cognitive impairment and Alzheimer’s dementia. Because circulating amyloid‐forming amylin triggers systemic hypoxia signaling, we sought to determine the relationship between circulating amylin levels and brain hypoxia markers in a transgenic rat model of amylin‐mediated neurological deficits.MethodTo assess amylin‐related brain hypoxia signaling, we performed a 16‐month longitudinal study in which rats that express amyloid‐forming human amylin in pancreatic β‐cells (HIP rats) were compared to wild type (WT) littermates that express non‐amyloidogenic rodent amylin (n=10 rats/group).ResultBlood amylin levels measured by ELISA were ∼2‐fold higher in 16‐month old HIP rats compared to WT littermates (P<0.01). The Thiovlavin T (Th T) fluorescence signal intensity in blood lysates from HIP rats increased compared to that in WT littermates indicating the presence of amyloid‐forming in the circulation in HIP rats (P<0.01). This was associated with higher amylin concentration in brain microvessel lysates (P<0.05) and amylin immunoreactivity signal intensity in brain sections. Plasma erythropoietin, a marker of systemic hypoxia was higher in HIP rats than in WT littermates (P<0.05), which correlated with brain accumulation of HIF‐2α (P<0.05) and HIF‐1α (P=0.07), and with altered mitochondrial DNA in brain tissue (P<0.05).ConclusionIncreased circulating levels of amyloid‐forming amylin promotes cerebrovascular amylin deposition leading to brain hypoxic‐ischemic injury. Future studies need to address functional effects of amylin‐induced brain hypoxia signaling in the brain.

18F]TRACK, a PET tracer for imaging of tropomyosin receptor kinases (Trk): efficient radiosynthesis and preliminary evaluation in the TgF344‐AD rat model of Alzheimer’s disease.

AbstractBackgroundA family of tropomyosin receptor kinases (TrkA/B/C) plays important roles in the regulation of neuronal differentiation, growth, and survival via their interactions with neurotrophins. A growing body of literature suggests the downregulation of Trk in many neurodegenerative conditions of the central nervous system (CNS), including Alzheimer’s disease (AD). We recently reported the development of [18F]TRACK, the PET tracers for in vivo imaging of TrkB and its first in‐human study. Here, we wish to report a fully automated GMP‐compliant radiosynthesis of [18F]TRACK at high radiochemical purity (RCP) and molar activity and its preliminary evaluation in wild‐type and a transgenic rat model of AD.MethodRadiosynthesis was performed using Scintomics GRP automated module via copper‐catalyzed 18F‐fluorination of the chiral boron pinacolate precursor. Brain imaging in wild‐type (n = 3) and TgF344‐AD rat models (n = 2) at 23 months was performed on microPET Concord R4 scanner.Result[18F]TRACK was synthesized with a 5.0±1.4 % radiochemical yield not corrected for decay (activity range 95–154 mCi) with >99% RCP and molar activities of 250 ± 75 GBq/µmol (n = 6). The synthesis was fully automated and all batches passed quality control procedures, including chemical purity, sterility and pyrogenicity. The tracer showed moderate brain uptake in wild‐type rats with the highest accumulation in the thalamus (SUVmax = 1.56 ± 0.28, 10 min post‐injection), consistent with the brain regional distribution in humans acquired in a parallel study. Preliminary data suggests lower tracer uptake in TgF344‐AD rat model of AD in thalamus, striatum and frontal cortex compared to wild‐type animals.ConclusionThis study demonstrated the feasibility of the GMP‐compliant production of [18F]TRACK in sufficient doses and molar activities for scanning multiple subjects per day or long range deliveries. Preliminary biodistribution studies showed moderate brain uptake of the tracer with preferential accumulation in the thalamus across species. These findings warrants further studies of Trk brain expression and its implication in neurodegenerative and psychiatric pathologies using [18F]TRACK.

An M1/sigma‐1 receptor agonist prevents cognitive deficits, reduces amyloid plaques and neuroinflammation in a transgenic rat model of Alzheimer’s amyloid pathology.

AbstractBackgroundAcetylcholinesterase inhibitors represent four out of six drugs for treating Alzheimer’s disease (AD), with time‐limited efficacy likely due to the progressive loss of cholinergic neurons. The selective allosteric M1 muscarinic and sigma‐1 receptor agonist Anavex 3‐71 (aka AF710B) [1] takes advantage of the fact that acetylcholine postsynaptic M1 muscarinic brain receptor levels remain unchanged in AD. Previously, Anavex 3‐71 treatment attenuated AD hallmarks in McGill‐R‐Thy1‐APP transgenic rats when administered at advanced stages of the AD‐like pathology [2]. However, preventive strategies instead of late treatment should be more effective in AD. Therefore, we tested whether Anavex 3‐71 administration during early amyloid pathology stages could prevent cognitive impairment, plaque deposition and neuroinflammation.MethodPre‐plaque, seven‐month‐old McGill‐R‐Thy1‐APP rats were subjected to daily oral administration of Anavex 3‐71 (10 µg/kg) for seven months. After one month of drug interruption (wash‐out), we performed Novel Object Recognition (NOR), Morris Water Maze (MWM) and Social Preference (SP) behavioural tests. Subsequently, brains were extracted, fixed and analyzed by histochemistry and immunohistochemistry.ResultMcGill‐APP rats had deficits in discriminating the novel object in the NOR and locating the hidden platform in the MWM compared to wild‐type littermates. Anavex 3‐71 restored the abilities of McGill‐APP rats in the NOR task to wild‐type levels and partially rescued the deficits in the MWM. In the SP task, Anavex 3‐71‐treated McGill‐APP rats exhibited diminished deficits in SP interaction time with a “stranger rat”. Treatment significantly reduced the number of mature Thioflavin‐S positive plaques in the subiculum compared to vehicle‐treated McGill‐APP rats and diminished the number of diffuse plaques in the cortex and hippocampus. Furthermore, Anavex 3‐71 treatment significantly reduced microglia and astrocyte recruitment towards CA1 hippocampal Aβ‐burdened neurons compared to untreated McGill‐APP rats.ConclusionThe long‐lasting effect of Anavex 3‐71 in preventing cognitive decline of McGill‐R‐Thy1‐APP rats, even after a wash‐out period would suggest some preventative, disease‐modifying, properties of the compound over the AD‐like amyloid pathology. This is supported by the attenuation of extracellular Aβ deposition and reduced glial cell recruitment towards hippocampal neurons. [1] Fisher et al., 2016; Neurodegenerative Diseases. https://doi.org/10.1159/000440864 [2] Hall et al., 2018; Alzheimer’s & Dementia. https://doi.org/10.1016/j.jalz.2017.11.009

Granulocyte‐Macrophage Colony‐Stimulating Factor Reduces Two Major Pathological Hallmarks of Alzheimer’s Disease and Astrogliosis in the TgF344‐AD Rat Model

AbstractBackgroundGranulocyte‐macrophage colony‐stimulating factor (GM‐CSF), a hematopoietic growth factor and the innate immune system modulator, has shown therapeutic and neuroprotective effects in animal models of Alzheimer’s disease (AD), Parkinson’s disease, Down syndrome, stroke, and normal aging. Recently, Potter et al. (2021) published the results of a Phase2 clinical trial in mild‐to‐moderate AD subjects showing that recombinant human GM‐CSF (sargramostim) treatment shifted both MMSE scores and plasma biomarkers of neurodegeneration toward normal. Cohen et al. (2013) developed a transgenic rat model (TgF344‐AD) that expresses the Swedish mutant human APP (APPsw) and mutant human presenilin 1 (PSEN1 deltaE9) genes that cause familial AD. In addition to cerebral amyloidosis, neuronal loss, and cognitive deficits, TgF344‐AD rats also show age‐dependent tauopathy including overexpression of phosphorylated tau and neurofibrillary tangle formation, which are not present in mouse models of AD. To evaluate the potential ability of GM‐CSF to reverse and/or prevent the full range of AD pathology, we treated ∼20‐month‐old TgF344‐AD rats with GM‐CSF or saline, and investigated AD pathology in the brain and retina. We also analyzed biomarkers in plasma using MSD and SIMOA neuroinflammation panels.MethodTgF344‐AD rats were injected subcutaneously with GM‐CSF (83.3ug/kg/day; 5days/week) or with saline (200ul/day) for 24 injections total over 32 days. On day 32, blood samples were collected, plasma samples were used for biomarkers assays, and brain and retinal tissues were processed for immunohistochemistry and immunoblotting.ResultTgF344‐AD rats treated with GM‐CSF showed a significant reduction in amyloid plaque deposition in the cortex and a strong trend towards reduced plaque deposition in the hippocampal CA region compared to saline‐treated TgF344‐AD rats. We also observed a trend of reduced phosphorylated tau expression in the cortex and hippocampal region in GM‐CSF‐treated TgF344‐AD rats. In addition, GM‐CSF treatment significantly reduced astrogliosis in the CA1 and dentate gyrus/hilus regions of TgF344‐AD rats.ConclusionAlthough experiments are ongoing, our findings indicate that GM‐CSF treatment rescues two major pathological hallmarks of AD and neuroinflammation (astrogliosis) in TgF344‐AD rats. Our results suggest that GM‐CSF/sargramostim/Leukine® may be a potential therapeutic not only for reducing amyloidosis, but also for reducing levels of hyperphosphorylated tau and other relevant pathological manifestations of AD.

Intronic enhancers of the human SNCA gene predominantly regulate its expression in brain in vivo.

Evidence from patients with Parkinson's disease (PD) and our previously reported α-synuclein (SNCA) transgenic rat model support the idea that increased SNCA protein is a substantial risk factor of PD pathogenesis. However, little is known about the transcription control of the human SNCA gene in the brain in vivo. Here, we identified that the DYT6 gene product THAP1 (THAP domain-containing apoptosis-associated protein 1) and its interaction partner CTCF (CCCTC-binding factor) act as transcription regulators of SNCA. THAP1 controls SNCA intronic enhancers' activities, while CTCF regulates its enhancer-promoter loop formation. The SNCA intronic enhancers present neurodevelopment-dependent activities and form enhancer clusters similar to "super-enhancers" in the brain, in which the PD-associated single-nucleotide polymorphisms are enriched. Deletion of the SNCA intronic enhancer clusters prevents the release of paused RNA polymerase II from its promoter and subsequently reduces its expression drastically in the brain, which may provide new therapeutic approaches to prevent its accumulation and thus related neurodegenerative diseases defined as synucleinopathies.

Use of multimodality imaging, histology, and treatment feasibility to characterize a transgenic Rag2-null rat model of glioblastoma.

Many drugs that show potential in animal models of glioblastoma (GBM) fail to translate to the clinic, contributing to a paucity of new therapeutic options. In addition, animal model development often includes histologic assessment, but multiparametric/multimodality imaging is rarely included despite increasing utilization in patient cancer management. This study developed an intracranial recurrent, drug-resistant, human-derived glioblastoma tumor in Sprague-Dawley Rag2-Rag2 tm1Hera knockout rat and was characterized both histologically and using multiparametric/multimodality neuroimaging. Hybrid 18F-fluoroethyltyrosine positron emission tomography and magnetic resonance imaging, including chemical exchange saturation transfer (18F-FET PET/CEST MRI), was performed for full tumor viability determination and characterization. Histological analysis demonstrated human-like GBM features of the intracranially implanted tumor, with rapid tumor cell proliferation (Ki67 positivity: 30.5 ± 7.8%) and neovascular heterogeneity (von Willebrand factor VIII:1.8 to 5.0% positivity). Early serial MRI followed by simultaneous 18F-FET PET/CEST MRI demonstrated consistent, predictable tumor growth, with exponential tumor growth most evident between days 35 and 49 post-implantation. In a second, larger cohort of rats, 18F-FET PET/CEST MRI was performed in mature tumors (day 49 post-implantation) for biomarker determination, followed by evaluation of single and combination therapy as part of the model development and validation. The mean percentage of the injected dose per mL of 18F-FET PET correlated with the mean %CEST (r = 0.67, P < 0.05), but there was also a qualitative difference in hot spot location within the tumor, indicating complementary information regarding the tumor cell demand for amino acids and tumor intracellular mobile phase protein levels. Finally, the use of this glioblastoma animal model for therapy assessment was validated by its increased overall survival after treatment with combination therapy (temozolomide and idasanutlin) (P < 0.001). Our findings hold promise for a more accurate tumor viability determination and novel therapy assessment in vivo in a recently developed, reproducible, intracranial, PDX GBM.

A Cre-driver rat model for anatomical and functional analysis of glucagon (Gcg)-expressing cells in the brain and periphery

ObjectiveThe glucagon gene (Gcg) encodes preproglucagon, which is cleaved to form glucagon-like peptide 1 (GLP1) and other mature signaling molecules implicated in metabolic functions. To date there are no transgenic rat models available for precise manipulation of GLP1-expressing cells in the brain and periphery. MethodsTo visualize and manipulate Gcg-expressing cells in rats, CRISPR/Cas9 was used to express iCre under control of the Gcg promoter. Gcg-Cre rats were bred with tdTomato reporter rats to tag Gcg-expressing cells. Cre-dependent AAVs and RNAscope in situ hybridization were used to evaluate the specificity of iCre expression by GLP1 neurons in the caudal nucleus of the solitary tract (cNTS) and intermediate reticular nucleus (IRt), and by intestinal and pancreatic secretory cells. Food intake was assessed in heterozygous (Het) Gcg-Cre rats after chemogenetic stimulation of cNTS GLP1 neurons expressing an excitatory DREADD. ResultsWhile genotype has minimal effect on body weight or composition in chow-fed Gcg-Cre rats, homozygous (Homo) rats have lower plasma glucose levels. In neonatal and adult Gcg-Cre/tdTom rats, reporter-labeled cells are present in the cNTS and IRt, and in additional brain regions (e.g., basolateral amygdala, piriform cortex) that lack detectable Gcg mRNA in adults but display transient developmental or persistently low Gcg expression. Compared to wildtype (WT) rats, hindbrain Gcg mRNA and GLP1 protein in brain and plasma are markedly reduced in Homo Gcg-Cre rats. Chemogenetic stimulation of cNTS GLP1 neurons reduced overnight chow intake in males but not females, the effect in males was blocked by antagonism of central GLP1 receptors, and hypophagia was enhanced when combined with a subthreshold dose of cholecystokinin-8 to stimulate gastrointestinal vagal afferents. ConclusionsGcg-Cre rats are a novel and valuable experimental tool for analyzing the development, anatomy, and function of Gcg-expressing cells in the brain and periphery. In addition, Homo Gcg-Cre rats are a unique model for assessing the role of Gcg-encoded proteins in glucose homeostasis and energy metabolism.

Probing changes in brain esterified oxylipin concentrations during the early stages of pathogenesis in Alzheimer’s Disease transgenic rats

Despite known pathological hallmarks of Alzheimer’s Disease (AD) including neuronal loss, gliosis (inflammation), beta-amyloid plaque deposition and neurofibrillary tangle accumulation in the brain, little is known about inflammation resolution in early AD pathogenesis. In the brain, inflammation and resolution pathways are mediated by free oxylipins which are mostly bound (i.e. esterified), and therefore must be released (i.e. become free) to exert bioactivity. Recently, we showed reductions in brain esterified pro-resolving oxylipins in a transgenic rat model of AD (TgF344-AD rat) at 15 months of age, suggesting deficits in the source and availability of free pro-resolving oxylipins. In the present study, we tested whether these changes are discernable earlier in the disease process, i.e., at age of 10 months. We observed significant reductions in esterified pro-resolving 8(9)-epoxyeicosatrienoic acid (8(9)-EpETrE), 13-hydroxyoctadecatrienoic acid (13-HOTrE) and 15-hydroxyeicosapentaenoic acid (15-HEPE) oxylipins, and in pro-inflammatory 13-hydroxy-octadecadienoic acid (13-HODE), 20-hydroxy-eicosatetraenoic acid (20-HETE), 15-deoxy-prostaglandin J2 (15-deoxy-PGJ2) and prostaglandin E2 (PGE2) oxylipins in male and/or female transgenic AD rats compared to wildtype controls. These findings point to a deficit in esterified pro-resolving lipid mediators in the early stages of AD, concident with. changes in esterified lipid mediators involved in promoting inflammation.

Early alterations in brain glucose metabolism and vascular function in a transgenic rat model of Alzheimer's disease.

Alteration in brain metabolism predates clinical onset of Alzheimer's Disease (AD). Realizing its potential as an early diagnostic marker, however, requires understanding how early AD metabolic dysregulation manifests on non-invasive brain imaging. We presently utilized magnetic resonance imaging and spectroscopy to map glucose and ketone metabolic profiles and image cerebrovascular function in a rat model of early stage AD - 9-month-old TgF344-AD (TgAD) rats - and their age-matched non-transgenic (nTg) littermates. Compared to the nTg rats, TgAD rats displayed attenuation in global cerebral and hippocampal vasoreactivity to hypercapnia, by 49±17% and 58±19%, respectively, while their functional hyperemia to somatosensory stimulation diminished by 69±5%. To assess brain glucose uptake, rats were fasted overnight and then challenged with an intravenous infusion of 2-deoxy-D-glucose (2DG). Compared to their non-transgenic littermates, TgAD rats exhibited 99±10% and 52±5% smaller glucose uptake in the entorhinal cortex and the hippocampus, respectively. Moreover, hippocampal glucose uptake reduction in male TgAD rats compared to the nTg was 54±36% greater than the reduction seen in female TgAD rats. TgAD rats also showed a 59±42% increase in total choline level in the hippocampus, suggesting increased membrane turnover. In combination with our earlier findings of impaired electrophysiological metrics at this early stage of AD pathology progression, our findings suggest that subtle neuronal function alterations that would be difficult to assess in a clinical population may be accompanied by MRI-detectable changes in brain glucose metabolism and cerebrovascular function.

Timapiprant, a prostaglandin D2 receptor antagonist, ameliorates pathology in a rat Alzheimer's model.

We investigated the relevance of the prostaglandin D2 pathway in Alzheimer's disease, because prostaglandin D2 is a major prostaglandin in the brain. Thus, its contribution to Alzheimer's disease merits attention, given the known impact of the prostaglandin E2 pathway in Alzheimer's disease. We used the TgF344-AD transgenic rat model because it exhibits age-dependent and progressive Alzheimer's disease pathology. Prostaglandin D2 levels in hippocampi of TgF344-AD and wild-type littermates were significantly higher than prostaglandin E2. Prostaglandin D2 signals through DP1 and DP2 receptors. Microglial DP1 receptors were more abundant and neuronal DP2 receptors were fewer in TgF344-AD than in wild-type rats. Expression of the major brain prostaglandin D2 synthase (lipocalin-type PGDS) was the highest among 33 genes involved in the prostaglandin D2 and prostaglandin E2 pathways. We treated a subset of rats (wild-type and TgF344-AD males) with timapiprant, a potent highly selective DP2 antagonist in development for allergic inflammation treatment. Timapiprant significantly mitigated Alzheimer's disease pathology and cognitive deficits in TgF344-AD males. Thus, selective DP2 antagonists have potential as therapeutics to treat Alzheimer's disease.

Neuropathology of the Basal Ganglia in SNCA Transgenic Rat Model of Parkinson's Disease: Involvement of Parvalbuminergic Interneurons and Glial-Derived Neurotropic Factor.

Parkinson’s disease (PD) is a neurodegenerative disease characterized by the accumulation of alpha-synuclein, encoded by the SNCA gene. The main neuropathological hallmark of PD is the degeneration of dopaminergic neurons leading to striatal dopamine depletion. Trophic support by a neurotrophin called glial-derived neurotrophic factor (GDNF) is also lacking in PD. We performed immunohistochemical studies to investigate neuropathological changes in the basal ganglia of a rat transgenic model of PD overexpressing alfa-synuclein. We observed that neuronal loss also occurs in the dorsolateral part of the striatum in the advanced stages of the disease. Moreover, along with the degeneration of the medium spiny projection neurons, we found a dramatic loss of parvalbumin interneurons. A marked decrease in GDNF, which is produced by parvalbumin interneurons, was observed in the striatum and in the substantia nigra of these animals. This confirmed the involvement of the striatum in the pathophysiology of PD and the importance of GDNF in maintaining the health of the substantia nigra.

Abstract P320: Transgenic Cre Drivers For Harvesting Podocytes And Endothelial Cells From Dahl Salt-sensitive Rats

Complex phenotypes such as blood pressure regulation involve multiple organ systems, tissues, and specific cell types. Transgenic rodent models with the expression of fluorescent proteins under the control of tissue- and cell-type-specific promoters are powerful tools for identifying and isolating specific cells but have been primarily limited to mice. Our long-term goal is to establish a Cre-driver rat resource for conditional and physiologically predictive rat models of human diseases. We developed two Cre-driver transgenic strains of rats (SS/PodocinCreERT2 and SS/VECadCre) capable of activating a second Cre-sensitive tdTomato reporter transgene integrated into the rat Rosa26 locus in the Dahl salt-sensitive hypertension model background. Cells with reporter activation were identified under fluorescence microscopy or harvested using fluorescence-activated cell sorting (FACS). A tamoxifen-inducible Cre transgene under the control of the Nphs2 (podocin) gene promoter was used to harvest fluorescently labeled podocytes after tamoxifen injection. A constitutive Cre under the control of the Cdh5 (VE-Cadherin) gene promoter was used for the isolation of endothelial cells. Correspondingly, tens to hundreds of thousands of podocytes or endothelial cells were isolated from a single kidney or mesenteric artery. Isolated cells were enriched for specific marker genes and could be used in downstream assays including single-cell transcriptional profiling. These new Cre/reporter dual transgenic rat models provide a much-needed resource for the biomedical research community to study the role of these cell types in a broad range of health and disease modeling studies.

Overexpression of cardiac‐specific IGF‐IIRα accelerates the development of liver dysfunction through STZ‐induced diabetic hepatocyte damage in transgenic rats

This study reports the effect of cardiac-specific insulin-like growth factor-II receptor α (IGF-IIRα) overexpression on the development of liver dysfunction in transgenic rats via STZ-induced diabetic hepatocyte damage. The cardio-hepatic syndrome comprises a number of heart and liver illnesses in which an acute or chronic disease in one organ can lead to acute or chronic disease in the other. However, the molecular mechanism involved in such a set of conditions is unclear. In this study, we developed a transgenic rat model with cardiac-specific overexpression of IGF-IIRα, which is a supplementary splicing variant of insulin-like growth factor-II receptor (IGF-IIR), expressed in pathological hearts, to investigate the relationship between late fetal gene expression in diabetic hearts and their influence on diabetic hepatopathy. STZ (55 mg/kg) was intraperitoneally delivered into IGF-IIR overexpressed transgenic (TG) and non-transgenic (NTG) animal models developed in Sprague-Dawley (SD) rats after an overnight fast. The relationship among IGF-IIRα overexpression and hepatocyte damages have been determined based on the complexity of damage in the liver. Our findings revealed that overexpression of the cardiac-specific IGF-IIRα enhances diabetes-induced morphological alterations and hepatic inflammation in the livers. The diabetic transgenic rats demonstrated the development of pathological conditions such as thick collagen fiber deposition, bridging fibrosis, and elevation of α-SMA and MMP1 related liver fibrosis mechanisms. Our data suggest that IGF-IIRα overexpression in the heart during a pathological state may worsen diabetic hepatopathy in rats.

Distinct Brain Proteomic Signatures in Cerebral Small Vessel Disease Rat Models of Hypertension and Cerebral Amyloid Angiopathy.

Cerebral small vessel diseases (CSVDs) are prominent contributors to vascular cognitive impairment and dementia and can arise from a range of etiologies. Cerebral amyloid angiopathy (CAA) and hypertension (HTN), both prevalent in the elderly population, lead to cerebral microhemorrhages, macrohemorrhages, and white matter damage. However, their respective underlying mechanisms and molecular events are poorly understood. Here, we show that the transgenic rat model of CAA type 1 (rTg-DI) exhibits perivascular inflammation that is lacking in the spontaneously hypertensive stroke-prone (SHR-SP) rat model of HTN. Alternatively, SHR-SP rats display notable dilation of arteriolar perivascular spaces. Comparative proteomics analysis revealed few shared altered proteins, with key proteins such as ANXA3, H2A, and HTRA1 unique to rTg-DI rats, and Nt5e, Flot-1 and Flot-2 unique to SHR-SP rats. Immunolabeling confirmed that upregulation of ANXA3, HTRA1, and neutrophil extracellular trap proteins were distinctly associated with rTg-DI rats. Pathway analysis predicted activation of TGF-β1 and TNFα in rTg-DI rat brain, while insulin signaling was reduced in the SHR-SP rat brain. Thus, we report divergent protein signatures associated with distinct cerebral vessel pathologies in the SHR-SP and rTg-DI rat models and provide new mechanistic insight into these different forms of CSVD.