We thank Drs. Xiaofang Wang and Katharina Hopp for their invaluable help with the animal models. This work has been supported by grants from the National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health (DK090728, DK058816).

Prima di avviare qualsiasi procedura con gli animali vivi, protocolli sperimentali dovranno essere approvati dalla cura degli animali e l&#39;uso comitato istituzionale (IACUC). 1. Configurazione Scanner <ol><li> Prima di iniziare, assicurarsi che il riscaldatore si trova in posizione OFF. </li><li> Selezionare il mini gradiente di imaging e 38 millimetri bobina RF e mini supporto imaging. </li><li> Nel foro centrale del supporto installare il gruppo a temperatura variabile. </li></ol> 2. Preparazione degli animali <ol><li> Per gli esperimenti di risonanza magnetica, ottenere l&#39;anestesia ottimali utilizzando isoflurano vaporizzato. Per l&#39;induzione dell&#39;anestesia, mettere animale in una camera di induzione foderato con un tessuto assorbente. Regolare il flussometro del vaporizzatore isoflurano a 2,0-2,5 L / min, e la isoflurano al 3% di ossigeno. </li><li> Rimuovere qualsiasi etichetta di metallo o altro oggetto metallico in questa fase. Applicare una pomata veterinario sugli occhi dell&#39;animale per prevenire la secchezza mentre sotto anestesia. </li><li> Una volta che l&#39;animaleha raggiunto il piano chirurgico di anestesia (cioè, perdita di recesso reflex a pizzico tep), posizionare l&#39;animale su un supporto con il suo naso inserito in un cono di naso. Impostare il flusso d&#39;aria anestesia nella sonda di 2,0-2,5 ml / min e la concentrazione isoflurano al 1,5-2,0% di ossigeno. Anestesia sarà consegnato attraverso il cono naso durante la procedura. Periodicamente regolare la concentrazione isoflurano a seconda dell&#39;età e del peso dell&#39;animale a mantenere un tasso di respirazione di ~ 40 bpm. </li><li> Utilizzare i titolari animali per garantire l&#39;animale al suo posto e impedire il movimento durante l&#39;esperimento MRI. Variare il tipo di supporto animale a seconda della regione del corpo da sottoporre a scansione. Nota: i titolari di misura dalla plastica di laboratorio (polipropilene, teflon, polistirolo, policarbonato) possono essere fatte per ospitare specifico esperimento e per adattarsi alle dimensioni degli animali (da topo neonato per 160 g di ratto). </li><li> Posizionare il termometro rettale nell&#39;animale per controllare la temperatura del corpo dell&#39;animale. Durante l&#39;experiment, tenere l&#39;animale a 35-37 ° C, in corrente di aria calda. Regolare la temperatura dell&#39;aria (30-38 ° C) e il flusso (1,200-2,000 l / h) sulla base del feedback della temperatura corporea dell&#39;animale. </li><li> Collegare un sensore di pressione respiratoria palloncino per addome dell&#39;animale per monitorare la frequenza respiratoria. </li><li> Fissare l&#39;animale al centro della bobina RF e posizionare accuratamente la bobina RF con l&#39;animale in MRI scanner. </li></ol> 3. MRI Experiment <ol><li> Tune e abbinare bobina RF prima di iniziare gli esperimenti per minimizzare la potenza RF utilizzato e per massimizzare il rapporto segnale-rumore. Per avviare l&#39;incontro / tuning: <ol><li> Aprire lo strumento di controllo spettrometro facendo clic sull&#39;icona strumenti. </li><li> Nello strumento di controllo spettrometro clic Acquisizione → Wobble. Una finestra Acq / Reco aprirà la visualizzazione della curva di oscillazione. </li><li> In alternativa, regolare i condensatori di sintonia e corrispondenza (utilizzando la messa a punto e di corrispondenza bacchette) a piccoli passi fino a quando la potenza RF riflessaè ridotto al minimo. L&#39;obiettivo è quello di vedere una curva con un minimo in corrispondenza dell&#39;asse verticale posizionato a zero sull&#39;asse orizzontale. </li><li> Quando la calibrazione della batteria è stato raggiunto con successo, premere il pulsante Stop nella finestra Acq / Reco. </li></ol></li><li> Acquisire le immagini esploratore nei tre piani ortogonali per creare immagini assiali, coronali e sagittali. Usare una sequenza di immagini veloce come Intra Porta veloce basso angolo di tiro (IG-FLASH) per acquisire le immagini da ricognizione 12. Utilizzare le immagini da ricognizione per impostare la geometria corretta per imaging effettivo. </li><li> A seconda degli obiettivi specifici di ricerca, selezionare la corretta sequenza di immagini e parametri e avviare la scansione con un semaforo. Questo calibrare canale RF, shim del magnete, impostato frequenza portante in risonanza per l&#39;acqua e regolare il guadagno del ricevitore, il tutto automaticamente. <ol><li> Per gli studi anatomici e T2 pesate, acquisire in fetta più 2D o in modalità 3D. Per ridurre il tempo per un dato esperimento risoluzione spaziale, mantenere il campo-di-view (FOV) più piccola possibile, ma abbastanza grande per evitare artefatti avvolgenti (2,56-3,2 cm). </li></ol></li><li> Mantenere il ciclo della sequenza selezionata leggermente più corto del ciclo respirazione animale dalla scelta corretta del tempo di ripetizione (TR) e / o il numero di fette. Ciò assicura che i dati vengono raccolti durante il periodo di quiete degli animali. <ol><li> Ad esempio, per le immagini addominali, mantenere la frequenza respiratoria dell&#39;animale a ~ 30 bpm; che è di circa 2.000 msec per respiro. Utilizzare un Turbo Rapid acquisizione con Enhancement Relax (RARE) Sequenza e acquisire 11-19 fette coronali, con TR / TE 1500/9 msec, fattore RARO 8 e (matrice 256 x 256, FOV 2,56 x 2,56 centimetri, spessore dello strato 0,75 millimetri) . Nota: Regolando il TR a 1.500 msec, e mantenendo la frequenza respiratoria dell&#39;animale ~ 30 bpm (2.000 msec per respiro), ci assicuriamo che i dati vengono raccolti durante il periodo di quiete degli animali. </li></ol></li><li> Dopo tutto acquisizione dell&#39;immagine è stata completata, posizionare l&#39;animale digitalizzatasul rilievo riscaldata e monitorare fino ambulatoriale. Dopo il recupero, rispedire l&#39;animale verso la gabbia e monitorati per almeno 1 ora prima di tornare alla struttura degli animali. </li></ol>

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The authors have nothing to disclose.

Questo manoscritto mostra la possibilità di utilizzare la risonanza magnetica UHF come strumento per la caratterizzazione fenotipica in vivo o il monitoraggio di farmaci in modelli di roditori per PKD. Descriviamo esperimenti fatti a 16,4 T con una vasta foro Avance III alta risoluzione spettrometro NMR dotata di accessori di imaging micro e mini. Lo spettrometro è stato guidato dal software di acquisizione ed elaborazione TopSpin2.0PV controllato da Paravision 5.1 software di ima...

La malattia del rene policistico (PKD) comprende un gruppo di malattie monogeniche caratterizzate dallo sviluppo di cisti renali. Tra loro ci sono autosomica dominante rene policistico (ADPKD) e autosomica recessiva rene policistico (ARPKD), che rappresentano i più comuni tipi 1,2. ADPKD, la forma più frequente di malattie ereditarie cistiche renali, si origina da mutazioni nei geni PKD1 o PKD2. Essa è caratterizzata da esordio tardivo, più cisti renali bilaterali, accompagnato da cisti extra-renale variabile, così come anomalie scheletriche cardiovascolari e muscolari. ARPKD, la maggior parte dei neonati che generalmente influenzano e bambini piccoli, è causata da mutazioni in PKHD1 ed è caratterizzata da reni ecogene dilatati e fibrosi epatica congenita 3. È importante sottolineare che, ADPKD è caratterizzata da eterogeneità, sia al gene (genica) e la mutazione (allelica) livelli, che si traduce in notevoli pla variabilità henotypic. Mutazioni nel gene PKD1 sono associati con grave quadro clinico (numerose cisti, la diagnosi precoce, l&#39;ipertensione, e ematuria), così come la progressione rapida stadio terminale della malattia renale (20 anni prima rispetto ai pazienti con mutazioni PKD2) 4. Malattia grave del fegato policistico (PLD) e anomalie vascolari possono essere associati a mutazioni in entrambi PKD1 e PKD2 5. La maggior parte delle complicanze renali di ADPKD ottengono prevalentemente in conseguenza dell&#39;espansione cisti con infiammazione e fibrosi associata. Sviluppo Cisti inizia in utero e continua attraverso il ciclo di vita del paziente. I reni normalmente mantengono la loro forma reniforme, anche se potevano raggiungere più di 20 volte il volume renale normale. La maggior parte dei pazienti presente distribuzione bilaterale di cisti renali, ma in alcuni casi particolari, cisti possono sviluppare in un modello unilaterale o asimmetrica. Un importante Challenge per nefrologi seguito i pazienti con ADPKD o terapie attuazione è la storia naturale della malattia. Durante la maggior parte del suo corso, la funzione renale rimane normale e quando la funzione renale inizia a diminuire, la maggior parte dei reni sono stati sostituiti da cisti. Quando le terapie sono implementati in fasi successive, è meno probabile che sia successo in quanto il paziente potrebbe aver già raggiunto un punto di non ritorno nella malattia renale cronica. Al contrario, quando le terapie vengono avviati nelle fasi iniziali, è difficile identificare una risposta basata esclusivamente sulla velocità di filtrazione glomerulare. Di conseguenza, il concetto di volume di rene come marker di progressione della malattia guadagnato l&#39;attenzione. Il Consorzio per Radiologic Imaging Studi di rene policistico Malattia (CRISP) studio ha dimostrato che nei pazienti affetti da ADPKD l&#39;incremento dei volumi di rene e cisti correla direttamente con deterioramento della funzione renale, sottolineando il potenziale di Volume totale del rene (TKV) come comemarcatore urrogate per la progressione della malattia 6,7. Di conseguenza, TKV è attualmente utilizzato come endpoint primario o secondario in più studi clinici per ADPKD 2,8,9. Più modelli murini comprese mutazioni spontanee e tecniche di ingegneria genetica hanno fatto luce sulla patogenesi di PKD 10,11. Modelli PKD1 o PKD2 (mutazioni in entrambe le PKD1 o PKD2) sono diventati più popolari, in quanto la malattia umana perfettamente imitare. Inoltre, i modelli di roditori con mutazioni in geni diversi geni PKD1 o PKD2 sono stati utilizzati come una piattaforma sperimentale di chiarire vie di segnalazione correlati alla malattia. Inoltre, molti di questi modelli sono stati utilizzati per testare potenziali terapie. Tuttavia, un fattore limitante in molti studi sui roditori per PKD è spesso la mancanza di metodi non invasivi efficienti per analizzare sequenzialmente i cambiamenti anatomici e funzionali nel rene. R magneticaesonance (MRI) è la tecnica di imaging attuale gold standard per seguire i pazienti ADPKD, fornire un eccellente contrasto dei tessuti molli e il dettaglio anatomico, e consentendo misure TKV. Anche se la risonanza magnetica è ben definito per l&#39;imaging anatomiche negli animali e nell&#39;uomo più grandi, di imaging piccoli roditori in vivo comporta sfide tecniche supplementari, dove la capacità di acquisire immagini ad alta risoluzione può limitarne l&#39;utilità. Con l&#39;introduzione di ultra-alto campo (UHF) MRI (7-16,4 T) e lo sviluppo di forti pendenze, è ora possibile ottenere una maggiore segnale-rumore rapporti e risoluzione spaziale di immagini MRI con una qualità diagnostica simile a quella ottenute negli uomini. Di conseguenza, l&#39;uso di UHF risonanza magnetica per imaging in vivo di piccoli modelli di roditori per PKD è diventato un potente strumento per i ricercatori.

<table><tbody><tr><td>AVANCEIII-700 (16.4 T)</td><td>Bruker</td><td>BH067206</td><td>Wide-bore two channel multinuclear spectrometer equipped with mini and micro-imaging accessories for &lt;em&gt;in vivo&lt;/em&gt; small rodent imaging</td></tr><tr><td>TopSpin2.0PV</td><td>Bruker</td><td>H9088TA2</td><td>Spectrometer processing software</td></tr><tr><td>Paravision 5.1</td><td>Bruker</td><td>T10314L5</td><td>Imaging sofware</td></tr><tr><td>VTU BVT 3000 digital</td><td>Bruker</td><td>W1101095</td><td>Temperature controller</td></tr></tbody></table>

use of ultra-high field mri in small rodent models of polycystic kidney disease for in vivo phenotyping and drug monitoring

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular mechanisms responsible for the disease, and are very convenient for rapid drug screening and testing of promising therapies. A limiting factor in these studies is often the lack of efficient non-invasive methods for sequentially analyzing the anatomical and functional changes in the kidney. Magnetic resonance imaging (MRI) is the current gold standard imaging technique to follow autosomal dominant polycystic kidney disease (ADPKD) patients, providing excellent soft tissue contrast and anatomic detail and allowing Total Kidney Volume (TKV) measurements.A major advantage of MRI in rodent models of PKD is the possibility for in vivo imaging allowing for longitudinal studies that use the same animal and therefore reducing the total number of animals required. In this manuscript, we will focus on using Ultra-high field (UHF) MRI to non-invasively acquire in vivo images of rodent models for PKD. The main goal of this work is to introduce the use of MRI as a tool for in vivo phenotypical characterization and drug monitoring in rodent models for PKD.

In questo manoscritto, ci proponiamo di mostrare l&#39;utilità di UHF MRI come strumento per la caratterizzazione fenotipica in vivo o il monitoraggio di farmaci in modelli di roditori per PKD e altre malattie renali. Tutti gli esperimenti erano parte di protocolli sperimentali approvati dal IACUC. In vivo fenotipizzazione di piccoli modelli di roditori per PKD utilizzo UHF MRI: Tutti gli studi di imaging sono stati eseguiti su animali vivi in ane...

Watch this Scientific Journal Video about Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring at JoVE.com

Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

The use of ultra-high field MRI as a non-invasive way to obtain phenotypic information of rodent models for polycystic kidney disease and to monitor interventions is described. Compared with the traditional histological approach, MRI images can be acquired in vivo, allowing for longitudinal follow-up.

L&#39;uso di ultra-alto campo MRI in piccolo roditore modelli di malattia del rene policistico per<em&gt; In Vivo</em&gt; Fenotipizzazione and Drug Monitoring

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular ...

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Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

11.4K Views.  Mayo Clinic. The use of ultra-high field MRI as a non-invasive way to obtain phenotypic information of rodent models for polycystic kidney disease and to monitor interventions is described. Compared with the traditional histological approach, MRI images can be acquired in vivo, allowing for longitudinal follow-up.

Video: Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

Utilizing High Resolution Ultrasound to Monitor Tumor Onset and Growth in Genetically Engineered Pancreatic Cancer Models

The LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre (KPC) mouse model represents an established and frequently used transgenic model to evaluate novel therapies in pancreatic cancer. Tumor onset is variable in the KPC model between 8 weeks and several months. Therefore, non-invasive imaging tools are required to screen for tumor onset and monitor for response to treatment. To address this issue, different approaches have emerged over the last years. High resolution ultrasound has major advantages such as non-invasiveness, fast session times and a high image resolution without radiation exposure. However, ultrasound in mice is not trivial and sufficient anatomical knowledge and practical skills are required to successfully perform high resolution ultrasound in preclinical pancreatic cancer models. With the following article, a detailed hands-on guide for abdominal ultrasound in murine models with a particular focus on endogenous pancreatic cancer models is presented. Furthermore, a summary of common mistakes and how to avoid them is provided.

This article describes the utilization of high-resolution ultrasound in genetically engineered pancreatic cancer mice. The primary aim is to provide a detailed instruction for detection and evaluation of endogenous pancreatic tumors.

Utilizzando ultrasuoni ad alta risoluzione per Monitor l'inizio del tumore e la crescita nei modelli geneticamente del cancro del pancreas

The LSL-KrasG12D/+; LSL-Trp53R172H/+; Pdx-1-Cre (KPC) mouse model represents an established and frequently used transgenic model to evaluate novel ...

Ricerca

Ricerca sul cancro

Multiple-mouse Neuroanatomical Magnetic Resonance Imaging

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and evaluating mouse models of human disease. MRI is an excellent modality for investigating genetically altered animals. It is capable of whole brain coverage, can be used in vivo, and provides multiple contrast mechanisms for investigating different aspects of neuranatomy and physiology. The advent of high-field scanners along with the ability to scan multiple mice simultaneously allows for rapid phenotyping of novel mutations.
Effective mouse MRI studies require attention to many aspects of experiment design. In this article, we will describe general methods to acquire quality images for mouse phenotyping using a system that images mice concurrently in shielded transmit/receive radio frequency (RF) coils in a common magnet (Bock et al., 2003). We focus particularly on anatomical phenotyping, an important and accessible application that has shown a high potential for impact in many mouse models at our imaging centre. Before we can provide the detailed steps to acquire such images, there are important practical considerations for both in vivo brain imaging (Dazai et al., 2004) and ex vivo brain imaging (Spring et al., 2007) that should be noted. These are discussed below.

Magnetic resonance imaging (MRI) has become an increasingly popular tool for examining the phenotype of genetically altered mice. This article illustrates the methods necessary to achieve high-throughput phenotyping of genetically altered mice using multiple-mouse MRI.

Multiple-topo Imaging neuroanatomici Risonanza Magnetica

The field of mouse phenotyping with magnetic resonance imaging (MRI) is rapidly growing, motivated by the need for improved tools for characterizing and ...

Neuroscienze

Early Detection of Drug-Induced Renal Hemodynamic Dysfunction Using Sonographic Technology in Rats

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is estimated to contribute to 19- 25% of all clinical cases of acute kidney injury (AKI) in critically ill patients. AKI detection has historically relied on metrics such as serum creatinine (sCr) or blood urea nitrogen (BUN) which are demonstrably inadequate in full assessment of nephrotoxicity in the early phase of renal dysfunction. Currently, there is no robust diagnostic method to accurately detect hemodynamic alteration in the early phase of AKI while such alterations might actually precede the rise in serum biomarker levels. Such early detection can help clinicians make an accurate diagnosis and help in in decision making for therapeutic strategy. Rats were treated with Cisplatin to induce AKI. Nephrotoxicity was assessed for six days using high-frequency sonography, sCr measurement and upon histopathology of kidney. Hemodynamic evaluation using 2D and Color-Doppler images were used to serially study nephrotoxicity in rats, using the sonography. Our data showed successful drug-induced kidney injury in adult rats by histological examination. Color-Doppler based sonographic assessment of AKI indicated that resistive-index (RI) and pulsatile-index (PI) were increased in the treatment group; and peak-systolic velocity (mm/s), end-diastolic velocity (mm/s) and velocity-time integral (VTI, mm) were decreased in renal arteries in the same group. Importantly, these hemodynamic changes evaluated by sonography preceded the rise of sCr levels. Sonography-based indices such as RI or PI can thus be useful predictive markers of declining renal function in rodents. From our sonography-based observations in the kidneys of rats that underwent AKI, we showed that these noninvasive hemodynamic measurements may consider as an accurate, sensitive and robust method in detecting early stage kidney dysfunction. This study also underscores the importance of ethical issues associated with animal use in research.

Early stage hemodynamic dysfunction is critical to the development of kidney disease. Yet, detection methodologies are limited. Recent advances in sonography provide a noninvasive, accurate option for early detection of kidney injury. This study outlines a step-by-step, sonographic methodology for detecting kidney dysfunction using a drug-induced nephrotoxicity rat model.

La diagnosi precoce del farmaco-indotta renale emodinamica erettile Utilizzando ecografica Technology di ratti

The kidney normally functions to maintain hemodynamic homeostasis and is a major site of damage caused by drug toxicity. Drug-induced nephrotoxicity is ...

Medicina

Transcutaneous Assessment of Renal Function in Conscious Rodents

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and time-consuming. In addition, serial blood or urine samples are required, with the associated stress for the experimental animals. A recent technique significantly reduces the investment in time and resources, minimizing the invasiveness and the animal stress, but being equally valid as the traditional approaches. The method measures transcutaneously renal function. Using an optical device and the exogenous renal marker fluorescein isothiocyanate (FITC)-sinistrin, this technique is capable of measuring the elimination kinetics of the marker through the skin. With neither blood nor urine samples nor the associated laboratory assays needed, the results of the transcutaneous measurement are almost instantaneously available. The method has been already validated in different species and successfully applied in several models of renal pathology. Moreover, due to its minimally invasive characteristics, it is suitable for sequential measurements within the same animal. Here is provided a detailed protocol to carry out the transcutaneous assessment of renal function in rodents.

Determination of glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional procedures to measure this parameter are cumbersome and require a large investment of time. Here we describe a faster and minimally invasive method to determinate GFR transcutaneously.

La valutazione transcutanea della funzione renale nei roditori Conscious

Glomerular filtration rate (GFR) is the gold standard to assess overall kidney function. However, traditional methods to evaluate GFR are cumbersome and ...

Assessment of Kidney Function in Mouse Models of Glomerular Disease

The use of murine models to mimic human kidney disease is becoming increasingly common. Our research is focused on the assessment of glomerular function in diabetic nephropathy and podocyte-specific VEGF-A knock-out mice; therefore, this protocol describes the full kidney work-up used in our lab to assess these mouse models of glomerular disease, enabling a vast amount of information regarding kidney and glomerular function to be obtained from a single mouse. In comparison to alternative methods presented in the literature to assess glomerular function, the use of the method outlined in this paper enables the glomerular phenotype to be fully evaluated from multiple aspects. By using this method, the researcher can determine the kidney phenotype of the model and assess the mechanism as to why the phenotype develops. This vital information on the mechanism of disease is required when examining potential therapeutic avenues in these models. The methods allow for detailed functional assessment of the glomerular filtration barrier through measurement of the urinary albumin creatinine ratio and individual glomerular water permeability, as well as both structural and ultra-structural examination using the Periodic Acid Schiff stain and electron microscopy. Furthermore, analysis of the genes dysregulated at the mRNA and protein level enables mechanistic analysis of glomerular function. This protocol outlines the generic but adaptable methods that can be applied to all mouse models of glomerular disease.

This protocol describes a full kidney work-up that should be carried out in mouse models of glomerular disease. The methods allow for detailed functional, structural, and mechanistic analysis of glomerular function, which can be applied to all mouse models of glomerular disease.

Valutazione della funzione del rene in modelli murini di malattia glomerulare

The use of murine models to mimic human kidney disease is becoming increasingly common. Our research is focused on the assessment of glomerular function ...

Use of 3D Robotic Ultrasound for In Vivo Analysis of Mouse Kidneys

Common modalities for in vivo imaging of rodents include positron emission tomography (PET), computed tomography (CT), magnetic resonance imaging (MRI), and ultrasound (US). Each method has limitations and advantages, including availability, ease of use, cost, size, and the use of ionizing radiation or magnetic fields. This protocol describes the use of 3D robotic US for in vivo imaging of rodent kidneys and heart, subsequent data analysis, and possible research applications. Practical applications of robotic US are the quantification of total kidney volume (TKV), as well as the measurement of cysts, tumors, and vasculature. Although the resolution is not as high as other modalities, robotic US allows for more practical high throughput data collection. Furthermore, using US M-mode imaging, cardiac function may be quantified. Since the kidneys receive 20%-25% of the cardiac output, assessing cardiac function is critical to the understanding of kidney physiology and pathophysiology.

Use of 3D Robotic Ultrasound for In Vivo Analysis of Mouse Kidneys

This protocol demonstrates robotic ultrasound (US) as a practical, cost-effective, and quick alternative to traditional non-invasive image modalities.

Uso di ultrasuoni robotici 3D per l'analisi in vivo dei reni di topo

Common modalities for in vivo imaging of rodents include positron emission tomography (PET), computed tomography (CT), magnetic resonance imaging (MRI), ...

Using 2-Photon Microscopy to Quantify the Effects of Chronic Unilateral Ureteral Obstruction on Glomerular Processes

Applying novel microscopy methods to suitable animal disease models to explore the dynamic physiology of the kidney remains a challenge. Rats with surface glomeruli provide a unique opportunity to investigate physiological and pathophysiological processes using intravital 2-photon microscopy. Quantification of glomerular capillary blood flow and vasoconstriction and dilatation in response to drugs, permeability, and inflammation are just some of the processes that can be studied. In addition, transgenic rats, i.e., podocytes labeled with fluorescent dyes and other molecular biomarker approaches, provide increased resolution to directly monitor and quantify protein-protein interactions and the effects of specific molecular alterations.
In mice, which lack surface glomeruli after four weeks of age, unilateral ureteral obstruction (UUO) for several weeks has been used to induce surface glomeruli. As this induction model does not allow for baseline studies, we quantified the effects of UUO on glomerular processes in the UUO model in Munich Wistar Fr&#246;mter (MWF) rats, which have surface glomeruli under physiologic conditions. The UUO model for five weeks or more induced significant alterations to gross renal morphology, the peritubular and glomerular microvasculature, as well as the structure and function of tubular epithelia. Glomerular and peritubular red blood cell (RBC) flow decreased significantly (p &#60; 0.01), probably due to the significant increase in the adherence of white blood cells (WBCs) within glomerular and peritubular capillaries. The glomerular sieving coefficient of albumin increased from 0.015 &#177; 0.002 in untreated MWFs to 0.045 &#177; 0.05 in 5-week-old UUO MWF rats. Twelve weeks of UUO resulted in further increases in surface glomerular density and glomerular sieving coefficient (GSC) for albumin. Fluorescent albumin filtered across the glomeruli was not reabsorbed by the proximal tubules. These data suggest that using UUO to induce surface glomeruli limits the ability to study and interpret normal glomerular processes and disease alterations.

Here, we present a protocol using 2-photon microscopy in Munich Wistar Fromter rats with surface glomeruli to quantifythe effects of prolonged ureteral obstruction on glomerular dynamics and function.

Utilizzo della microscopia a 2 fotoni per quantificare gli effetti dell'ostruzione ureterale unilaterale cronica sui processi glomerulari

Applying novel microscopy methods to suitable animal disease models to explore the dynamic physiology of the kidney remains a challenge. Rats with surface ...

High-Resolution Cardiac Positron Emission Tomography/Computed Tomography for Small Animals

Positron emission tomography (PET) and computed tomography (CT) are among the most employed diagnostic imaging techniques, and both serve in understanding cardiac function and metabolism. In preclinical research, dedicated scanners with high sensitivity and high spatio-temporal resolution are employed, designed to cope with the demanding technological requirements posed by the small heart size and very high heart rates of mice and rats. In this paper, a bimodal cardiac PET/CT imaging protocol for experimental mouse and/or rat models of cardiac diseases is described, from animal preparation and image acquisition and reconstruction to image processing and visualization.
In particular, the 18F-labeled fluorodeoxyglucose ([18F]FDG)-PET scan allows for the measurement and visualization of glucose metabolism in the different segments of the left ventricle (LV). Polar maps are convenient tools to display this information. The CT part consists of a time-resolved 3D reconstruction of the entire heart (4D-CT) using retrospective gating without electrocardiography (ECG) leads, allowing the morphofunctional evaluation of the LV and the subsequent quantification of the most important cardiac function parameters, such as ejection fraction (EF) and stroke volume (SV). Using an integrated PET/CT scanner, this protocol can be executed within the same anesthesia induction without the need to reposition the animal between different scanners. Hence, PET/CT can be seen as a comprehensive tool for the morphofunctional and metabolic evaluation of the heart in several small animal models of cardiac diseases.

Here, we present an experimental imaging protocol for the quantification of cardiac function and morphology using high-resolution positron emission tomography/computed tomography for small animals. Both mice and rats are considered, discussing the different requirements of computed tomography contrast agents for the two species.

Tomografia cardiaca ad emissione di positroni ad alta risoluzione/tomografia computerizzata per piccoli animali

Positron emission tomography (PET) and computed tomography (CT) are among the most employed diagnostic imaging techniques, and both serve in understanding ...

Micropuncture of Bowman's Space in Mice Facilitated by 2 Photon Microscopy

Renal micropuncture and renal 2-photon imaging are seminal techniques in renal physiology. However, micropuncture is limited by dependence on conventional microscopy to surface nephron features, and 2-photon studies are limited in that interventions can only be assessed at the organ, rather than the nephron level. In particular, micropuncture studies of the glomeruli of mice have been challenged by the paucity of surface glomeruli in mice. To address this limitation in order to pursue studies of aspirate from Bowman&#39;s space in mouse physiologic models, we developed 2-photon glomerular micropuncture. We present a novel surgical preparation that allows lateral access to the kidney while preserving the required vertical imaging column for 2-photon microscopy. Administration of high molecular weight fluorescein isothiocyanate (FITC)-dextran is used to render the renal vasculature and therefore glomeruli visible for 2-photon imaging. A quantum dot-coated pipette is then introduced under stereotactic guidance to a glomerulus selected from the several to many which may be visualized within the imaging window. In this protocol, we provide details of the preparation, materials, and methods necessary to carry out the procedure. This technique facilitates previously-impossible physiologic study of the kidney, including recovery of filtrate from Bowman&#39;s space and all segments of the nephron within the imaging depth limit, about 100 &#181;m below the renal capsule. Pressure, charge and flow may all be measured using the introduced pipette. Here, we provide representative data from liquid chromatography/mass spectrometry performed on aspirate from Bowman&#39;s space. We expect this technique to have wide applicability in renal physiologic investigation.

We present use of 2-photon microscopy to place a micropipette within Bowman&#39;s urinary space in mice, combining 2 foundational techniques of renal physiology. Use of 2-photon microscopy overcomes critical limitations of conventional microscopy for micropuncture renal physiology studies.

Micropuntura dello spazio di Bowman in topi nei paraggi 2 Photon microscopia

Renal micropuncture and renal 2-photon imaging are seminal techniques in renal physiology. However, micropuncture is limited by dependence on conventional ...

Biologia

Ultrafast Doppler Ultrasound for Hepatic Microvasculature Analysis in Mice

Imaging and Quantification of the Hepatic Vasculature of Mice Using Ultrafast Doppler Ultrasound

Non-invasive in vivo imaging of the vasculature is a powerful tool for studying disease mechanisms in rodents. To achieve high sensitivity imaging of the microvasculature using Doppler ultrasound methods, imaging modalities employing the concept of ultrafast imaging are preferred. By increasing the frame rate of the ultrasound scanner to thousands of frames per second, it becomes possible to improve the sensitivity of the blood flow down to 2 mm/s and to obtain functional information about the microcirculation in comparison to a sensitivity of around 1 cm/s in conventional Doppler modes. While Ultrafast Doppler ultrasound (UFUS) imaging has become adopted in neuroscience, where it can capture brain activity through neurovascular coupling, it presents greater challenges when imaging the vasculature of abdominal organs due to larger motions linked to breathing. The liver, positioned anatomically under the diaphragm, is particularly susceptible to out-of-plane movement and oscillating respiratory motion. These artifacts not only adversely affect Doppler imaging but also complicate the anatomical analysis of vascular structures and the computation of vascular parameters. Here, we present a qualitative and quantitative imaging analysis of the hepatic vasculature in mice by UFUS. We identify major anatomical vascular structures and provide graphical illustrations of the hepatic macroscopical anatomy, comparing it to an in-depth anatomical assessment of the hepatic vasculature based on Doppler readouts. Additionally, we have developed a quantification protocol for robust measurements of hepatic blood volume of the microvasculature over time. To contemplate further research, qualitative vascular analysis provides a comprehensive overview and suggests a standardized terminology for researchers working with mouse models of liver disease. Furthermore, it offers the opportunity to apply ultrasound as a non-invasive complementary method to inspect hepatic vascular defects in vivo and measure functional microvascular alterations deep within the organ before unraveling blood vessel anomalies at the micron scale levels using ex vivo staining on tissue sections.

Ultrafast Doppler ultrasound (UFUS) with high spatial resolution (100 &#956;m) and sensitivity represents a suitable noninvasive imaging modality for obtaining a comprehensive qualitative overview of the hepatic vasculature. UFUS also facilitates quantitative measurements of the microvasculature, aiming to enhance our understanding of vascular disease mechanisms.

Non-invasive in vivo imaging of the vasculature is a powerful tool for studying disease mechanisms in rodents. To achieve high sensitivity imaging of the ...

L'uso di ultra-alto campo MRI in piccolo roditore modelli di malattia del rene policistico per

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