Recombinant human bone morphogenic protein-2 (rhBMP-2) use in spinal fusion is limited by dose-dependent complications. Peptide amphiphile (PA) supramolecular polymers presenting a BMP-2-binding epitope have previously been developed to reduce the rhBMP-2 dose required for successful fusion. We evaluated PA implant biodegradation and tissue clearance in a rat posterolateral spinal fusion model as a prerequisite to clinical safety studies. Twenty-three female Sprague-Dawley rats underwent L4-L5 fusion with gadolinium (Gd)-labeled PA implants. Longitudinal magnetic resonance imaging (MRI) was performed up to 13 weeks postoperatively, while the spine and filter organs were harvested for inductively coupled plasma mass spectrometry (ICP-MS) quantification of Gd at multiple time points. Gd concentration at the fusion site decreased from 71% of maximum to 19.5% at 13 weeks, and MRI showed a complete loss of Gd signal enhancement by 8 weeks. In peripheral organs, peak Gd accumulation was 3% in the liver at 4 weeks, declining to 1.4% at 13 weeks, while Gd remained below 0.05% in the spleen, lung, and blood at all time points. These data indicate PA implant localization, with robust degradation and clearance and minimal off-target accumulation, supporting its translational potential for spinal fusion applications.

Miniaturized implantable optoelectronic technologies for in vivo biomedical applications are gaining interest but require strict thermal management for safe operation. Here, we introduce a comprehensive framework combining analytical solutions and numerical modeling to estimate and manage thermal effects of optoelectronic devices. We propose Green’s functions to analytically solve temperature distributions in tissue from a point source with coupled thermal-optical power, capturing the influence of critical tissue properties and spatiotemporal parameters. Integrating the Green’s function derives temperature distributions for sources with definable geometries. Numerical modeling defines scaling factors to account for variations in radiation patterns and material designs, enabling direct performance comparisons across systems. Guided by this framework, iterative optimization of a filamentary optogenetic probe for deep brain stimulation significantly reduces thermal loads while preserving typical behaviors in freely moving mice. Experimental validation through in vitro and in vivo characterization demonstrates scalable strategies to overcome thermal challenges in advanced bio-optoelectronic systems.

Precise spatial regulation of site-specific DNA recombination (SSR) in vivo remains a challenge due to limited tunability of current platforms. Here, we present an optogenetic approach that overcome these limitations by employing engineered light-regulated recombinase E-LightR-Cre and tunable wireless implantable optoelectronic devices. E-LightR-Cre meets the key criteria for spatial regulation of SSR in vivo , showing no detectable activity in the dark, while demonstrating robust activation upon blue-light illumination. To achieve local E-LightR-Cre activation in murine lungs, we developed wireless, fully-implantable optoelectronic devices enabling focal illumination with no discernible organ damage. By modulating illumination intensity and duration, we can control the size of the activated area. Local expression of oncogenic KRas-G12D in a photoactivated subpopulation of cells in vitro revealed rapid reprogramming of the mutant expressing cells and their non-activated neighbors. Light-guided activation of E-LightR-Cre in mouse lungs resulted in focal expression of a reporter gene and allowed us to induce local formation of oncogenic lesions in vivo .

In chemotherapeutic treatments, while cancer cells are the primary target, cytotoxic side effects are an important consideration. In the current study, we applied an in vivo imaging tool for characterizing chemotherapeutic response in a preclinical setting. The study focused on simultaneously examining the tumor and tissue response as a result of treatment with bortezomib, a mainstay proteasome inhibitor for treating multiple myeloma, in a preclinical model. OPM-2 tumor-bearing SCID-beige mice were designated as control or treated with bortezomib (1 mg/kg, i.v., every 4 days) (n = 8 per group). 99mTc-duramycin SPECT/CT whole-body scans were acquired 2 days before treatment as baseline and at days 1, 3 and 5 after treatment. Radioactivity uptake in tissues and organs was determined and quantitatively compared between control and bortezomib-treated group at each of the time points. Based on the imaging data, separate groups of tumor-bearing mice (n = 3 each) were included as control and bortezomib treated and the tissues were collected on day 5 for histopathology. In vivo imaging data identified significantly elevated 99mTc-duramycin uptake in the tumor, particularly in tumoral periphery. This was accompanied with signal changes in multiple organs and tissues including the adipose tissue, major bones, abdominal regions, spleen and testes. The imaging findings were consistent with known cytotoxic side effects of bortezomib and were supported by histopathology. The outcome of the study demonstrated potential utilities of the technology by enabling timely determination of the efficacy of anticancer treatments and the effect on collateral tissues as a result of systemic cytotoxic treatment.

Because large brains are energetically expensive, they are associated with metabolic traits that facilitate energy availability across vertebrates. However, the biological underpinnings driving these traits are not known. Given its role in regulating host metabolism in disease studies, we hypothesized that the gut microbiome contributes to variation in normal cross-vertebrate species differences in metabolism, including those associated with the brain's energetic requirements. By inoculating germ-free mice with the gut microbiota (GM) of three primate species - two with relatively larger brains and one with a smaller brain - we demonstrated that the GM of larger-brained primates shifts host metabolism towards energy use and production, while that of smaller-brained primates stimulates energy storage in adipose tissues. Our findings establish a causal role of the GM in normal cross-host species differences in metabolism associated with relative brain size and suggest that the GM may have been an important facilitator of metabolic changes during human evolution that supported encephalization.

Comprehensive, continuous quantitative monitoring of intricately orchestrated physiological processes and behavioral states in living organisms can yield essential data for elucidating the function of neural circuits under healthy and diseased conditions, for defining the effects of potential drugs and treatments, and for tracking disease progression and recovery. Here, we report a wireless, battery-free implantable device and a set of associated algorithms that enable continuous, multiparametric physio-behavioral monitoring in freely behaving small animals and interacting groups. Through advanced analytics approaches applied to mechano-acoustic signals of diverse body processes, the device yields heart rate, respiratory rate, physical activity, temperature, and behavioral states. Demonstrations in pharmacological, locomotor, and acute and social stress tests and in optogenetic studies offer unique insights into the coordination of physio-behavioral characteristics associated with healthy and perturbed states. This technology has broad utility in neuroscience, physiology, behavior, and other areas that rely on studies of freely moving, small animal models.

Background The goal was to determine the feasibility of mapping the injured‐but‐not‐infarcted myocardium using 99m Tc‐duramycin in the postischemic heart, with spatial information for its characterization as a pathophysiologically intermediate tissue, which is neither normal nor infarcted. Methods and Results Coronary occlusion was conducted in Sprague Dawley rats with preconditioning and 30‐minute ligation. In vivo single‐photon emission computed tomography was acquired after 3 hours (n=6) using 99m Tc‐duramycin, a phosphatidylethanolamine‐specific radiopharmaceutical. The 99m Tc‐duramycin + areas were compared with infarct and area‐at‐risk (n=8). Cardiomyocytes and endothelial cells were isolated for gene expression profiling. Cardiac function was measured with echocardiography (n=6) at 4 weeks. In vivo imaging with 99m Tc‐duramycin identified the infarct (3.9±2.4% of the left ventricle and an extensive area 23.7±2.2% of the left ventricle) with diffuse signal outside the infarct, which is pathologically between normal and infarcted (apoptosis 1.8±1.6, 8.9±4.2, 13.6±3.8%; VCAM‐1 [vascular cell adhesion molecule 1] 3.2±0.8, 9.8±4.1, 15.9±4.2/mm 2 ; tyrosine hydroxylase 14.9±2.8, 8.6±4.4, 5.6±2.2/mm 2 ), with heterogeneous changes including scattered micronecrosis, wavy myofibrils, hydropic change, and glycogen accumulation. The 99m Tc‐duramycin + tissue is quantitatively smaller than the area‐at‐risk (26.7% versus 34.4% of the left ventricle, P =0.008). Compared with infarct, gene expression in the 99m Tc‐duramycin + –noninfarct tissue indicated a greater prosurvival ratio (BCL2/BAX [B‐cell lymphoma 2/BCL2‐associated X] 7.8 versus 5.7 [cardiomyocytes], 3.7 versus 3.2 [endothelial]), and an upregulation of ion channels in electrophysiology. There was decreased contractility at 4 weeks (regional fractional shortening −8.6%, P <0.05; circumferential strain −52.9%, P <0.05). Conclusions The injured‐but‐not‐infarcted tissue, being an intermediate zone between normal and infarct, is mapped in vivo using phosphatidylethanolamine‐based imaging. The intermediate zone contributes significantly to cardiac dysfunction.

BACKGROUND CONTEXT Current methods of spinal fusion exhibit significant rates of pseudoarthrosis (failed fusion) that are associated with significant morbidity. The addition of bone morphogenic protein (rhBMP-2) enhances spine fusion rates, but there are well established side effects. To mitigate these risks, implants consisting of peptide amphiphile (PA) nanofibers containing a BMP-2 binding epitope have been developed that potentiate BMP-2 activity, reducing the dose of rhBMP-2 required to elicit successful fusion in a pre-clinical model. PAs are composed of lipids and amino acids that self-assemble into cylindrical fibers in a structure analogous to an extracellular matrix of collagen fibrils. In moving towards clinical trials, a thorough understanding of the biodegradation rate, tissue distribution, and clearance are needed. This study sought to quantify the degradation rate and evaluate distribution of the material in the pre-clinical setting of posterolateral spinal fusion in rats. PURPOSE In moving towards clinical trials, a thorough understanding of the biodegradation rate, tissue distribution, and clearance are needed. This study sought to quantify the degradation rate and evaluate distribution of the material in the pre-clinical setting of posterolateral spinal fusion in rats. STUDY DESIGN/SETTING N/A PATIENT SAMPLE N/A OUTCOME MEASURES N/A METHODS The study was approved by the Northwestern University Institutional Animal Care and Use Committee. Twenty-three female Sprague-Dawley rats, aged 12-16 weeks underwent L4-L5 posterolateral fusion with bilateral placement of implants composed of porous collagen microparticles, gadolinium (Gd)-labeled nanofibers containing a BMP-2 binding epitope, and low dose rhBMP-2. Magnetic resonance imaging (MRI) was performed postoperatively at increasing time points out to 13 weeks, followed by tissue harvest (spine, blood, kidney, liver, lung, and spleen) for inductively coupled plasmonic mass spectroscopy (ICP-MS)-based quantification of Gd (N=3/timepoint). Three animals underwent longitudinal imaging until the 13-week study endpoint. The Gd detected in harvested tissues at each timepoint was calculated as a percent of the Gd in the original implant. Additionally, two spines were harvested at 10 weeks and one at 13 weeks for manual palpation-based fusion scoring and high-resolution microcomputed tomographic imaging (μCT) for visualization of the fusion mass. RESULTS Gd signal decreased gradually in the spine fusion site, from 71% of the presurgical implant concentration at 4 hours postoperatively, to 19% by 13 weeks. Among the peripheral organs, the highest accumulation of Gd was 3% of the presurgical implant concentration in the liver at 4 weeks, which declined to 1.4% at 13 weeks. Gd accumulation in the kidney peaked at 4 weeks (1.6%), declining to 0.11% at 13 weeks. For the duration of the study, Gd remained below 0.05% in the spleen, 0.03% in the lung, and 0.01% in the blood. MRI corroborates these findings with a notable loss of Gd signal at 8-weeks and complete loss of signal at the 13-week timepoint. The spines evaluated by manual palpation and μCT all showed bilateral fusion. CONCLUSIONS This study aimed to understand the degradation and biodistribution of a PA implant in a rat spinal fusion model. ICP-MS showed that the Gd-tagged implant remained localized in the surgical site, with limited accumulation in peripheral organs during degradation. By 13 weeks, only 19.5% of the initial Gd was detected locally in the spine. This was corroborated by lack of signal on MRI, supporting robust clearance after degradation. We posit that the Gd nadir was 19.5% due to Gd-PA nanofiber incorporation into the bony fusion mass. The Gd concentration in many of the clearance organs (blood, spleen, and lung) remained less than 0.1% for the duration of the study. This work shows timely degradation of this amino acid-based spinal fusion implant, with limited accumulation in peripheral organs and no change in spinal fusion efficacy due to the Gd label. This study provides new insight regarding the degradation and subsequent biodistribution of a peptide amphiphile-based implant material designed for bone regeneration. Our findings indicate that the PA implant remains well localized after placement at the L4-L5 transverse processes, without significant peripheral accumulation. FDA Device/Drug Status This abstract does not discuss or include any applicable devices or drugs.

Critical limb ischemia (CLI) presents a significant clinical challenge, leading to tissue ischemia and potentially resulting in limb necrosis or amputation. Cell-based regenerative therapies offer promise for improving outcomes in CLI, but their effectiveness is often limited by poor cell survival and engraftment. This study hypothesized that a thermo-responsive polymer, poly(polyethylene glycol citrate-co-N-isopropylacrylamide) (PPCN), combined with pro-survival bioactive peptides, can create a protective microenvironment to improve endothelial cell survival and function after their delivery. Through in vitro and in vivo experiments, laminin-derived peptide A5G81 and vascular endothelial growth factor (VEGF)-derived peptide QK are identified as effective in promoting endothelial cell spreading, proliferation, and prolonged survival. PPCN's viscoelastic properties protected against shear stress during injection, while the peptides supported endothelial cell behavior through distinct molecular pathways. Importantly, delivery of endothelial cells with PPCN-A5G81 and PPCN-QK in a murine hindlimb ischemia model resulted in significant improvements in limb perfusion, tissue preservation, and functional outcomes compared to controls. Additionally, this approach enhanced skeletal muscle remodeling following ischemic injury. This innovative biomaterial platform represents a versatile solution for addressing cell survival challenges and advancing regenerative therapies in CLI and other ischemic conditions.

The absence of clinically applicable imaging techniques for continuous monitoring of transplanted cells poses a significant obstacle to the clinical translation of stem cell-based therapies for vascular regeneration. This study aims to optimize a clinically applicable, non-invasive imaging technique to longitudinally monitor vascular endothelial cells (ECs) for vascular regeneration in peripheral artery disease (PAD). Human induced pluripotent stem cells (HiPSCs) were employed to generate ECs (HiPSC-ECs). Lentiviral vectors encoding human sodium iodide symporter (hNIS) and enhanced green fluorescent protein (eGFP) genes were introduced to HiPSCs and HiPSC-ECs at varying multiplicities of infection (MOI). Through a combination of fluorescence microscopy and flow cytometry, an optimized transduction technique for introducing hNIS-eGFP into HiPSC-ECs was established. Subsequently, single-photon emission computed tomography (SPECT) was utilized for imaging of the transduced cells in vitro and in vivo after transplantation into the gastrocnemius muscle of nude mice. Lentiviral transduction resulted in sustained co-expression of hNIS and eGFP in HiPSC-ECs when transduced post-endothelial differentiation. An optimal MOI of five yielded over 90

Introduction: Critical limb ischemia (CLI) causes severe morbidity and mortality with limited treatment options. Cell-based therapy has potential to advance treatment outcomes but faces challenges in cell survival and engraftment during delivery. We hypothesize that an antioxidant niche that promotes cell adhesion and spreading will improve cell engraftment and survival rate, with the goal of limb salvage through improved perfusion. Methods: Thermoresponsive poly(polyethylene glycol citrate-co-N-isopropylacrylamide) (PPCN) was synthesized and conjugated with laminin-derived (A5G81) or VEGF-mimic (QK) peptides. Human umbilical vein endothelial cells (HUVECs) expressing human sodium iodine symporter (hNIS) were mixed with materials and intramuscularly injected into nude mice hindlimbs and monitored with Single Photon Emission Computed Tomography with Computed Tomography (SPECT/CT) for cell survival. Next, a hindlimb ischemia model was used to assess the efficacy of cell delivery by evaluating limb perfusion, tissue loss, functional impairment and histological examination. Results: Cells delivered with PBS or PPCN demonstrated poor cell survival, with over 90% cell loss by day 7 ( Figure 1a ). In contrast, cells delivered with PPCN-A5G81 and PPCN-QK exhibited robust SPECT signal levels for up to four weeks ( Figure 1a,b ). In the hindlimb ischemia model, materials or cells alone did not demonstrate improvement in any of the parameters over the PBS control. However, cells delivered with PPCN-A5G81 and PPCN-QK showed significantly higher blood perfusion ( Figure 1c-e ), with improved tissue salvage and motor functions ( Figure 1f-i ) for both male and female mice. Conclusion: PPCN modified with A5G81 and QK promotes cell spreading, proliferation, survival, and tissue regeneration, with potential for more effective cell-based therapies for CLI.

BACKGROUND:The pathology in Duchenne muscular dystrophy (DMD) is characterized by degenerating muscle fibers, inflammation, fibro-fatty infiltrate, and edema, and these pathological processes replace normal healthy muscle tissue. The mdx mouse model is one of the most commonly used preclinical models to study DMD. Mounting evidence has emerged illustrating that muscle disease progression varies considerably in mdx mice, with inter-animal differences as well as intra-muscular differences in pathology in individual mdx mice. This variation is important to consider when conducting assessments of drug efficacy and in longitudinal studies. We developed a magnetic resonance imaging (MRI) segmentation and analysis pipeline to rapidly and non-invasively measure the severity of muscle disease in mdx mice. METHODS:Wildtype and mdx mice were imaged with MRI and T2 maps were obtained axially across the hindlimbs. A neural network was trained to rapidly and semi-automatically segment the muscle tissue, and the distribution of resulting T2 values was analyzed. Interdecile range and Pearson Skew were identified as biomarkers to quickly and accurately estimate muscle disease severity in mice. RESULTS:The semiautomated segmentation tool reduced image processing time approximately tenfold. Measures of Pearson skew and interdecile range based on that segmentation were repeatable and reflected muscle disease severity in healthy wildtype and diseased mdx mice based on both qualitative observation of images and correlation with Evans blue dye uptake. CONCLUSION:Use of this rapid, non-invasive, semi-automated MR image segmentation and analysis pipeline has the potential to transform preclinical studies, allowing for pre-screening of dystrophic mice prior to study enrollment to ensure more uniform muscle disease pathology across treatment groups, improving study outcomes.

The pathology in Duchenne muscular dystrophy (DMD) is characterized by degenerating muscle fibers, inflammation, fibro-fatty infiltrate, and edema, and these pathological processes replace normal healthy muscle tissue. The mdx mouse model is one of the most commonly used preclinical models to study DMD. Mounting evidence has emerged illustrating that muscle disease progression varies considerably in mdx mice, with inter-animal differences as well as intra-muscular differences in pathology in individual mdx mice. This variation is important to consider when conducting assessments of drug efficacy and in longitudinal studies. Magnetic resonance imaging (MRI) is a non-invasive method that can be used qualitatively or quantitatively to measure muscle disease progression in the clinic and in preclinical models. Although MR imaging is highly sensitive, image acquisition and analysis can be time intensive. The purpose of this study was to develop a semi-automated muscle segmentation and quantitation pipeline that can quickly and accurately estimate muscle disease severity in mice. Herein, we show that the newly developed segmentation tool accurately divides muscle. We show that measures of skew and interdecile range based on segmentation sufficiently estimate muscle disease severity in healthy wildtype and diseased mdx mice. Moreover, the semi-automated pipeline reduced analysis time by nearly 10-fold. Use of this rapid, non-invasive, semi-automated MR imaging and analysis pipeline has the potential to transform preclinical studies, allowing for pre-screening of dystrophic mice prior to study enrollment to ensure more uniform muscle disease pathology across treatment groups, improving study outcomes.

Fully implantable wireless systems for the recording and modulation of neural circuits that do not require physical tethers or batteries allow for studies that demand the use of unconstrained and freely behaving animals in isolation or in social groups. Moreover, feedback-control algorithms that can be executed within such devices without the need for remote computing eliminate virtual tethers and any associated latencies. Here we report a wireless and battery-less technology of this type, implanted subdermally along the back of freely moving small animals, for the autonomous recording of electroencephalograms, electromyograms and body temperature, and for closed-loop neuromodulation via optogenetics and pharmacology. The device incorporates a system-on-a-chip with Bluetooth Low Energy for data transmission and a compressed deep-learning module for autonomous operation, that offers neurorecording capabilities matching those of gold-standard wired systems. We also show the use of the implant in studies of sleep-wake regulation and for the programmable closed-loop pharmacological suppression of epileptic seizures via feedback from electroencephalography. The technology can support a broader range of applications in neuroscience and in biomedical research with small animals.

Continuous, real-time monitoring of perfusion after microsurgical free tissue transfer or solid organ allotransplantation procedures can facilitate early diagnosis of and intervention for anastomotic thrombosis. Current technologies including Doppler systems, cutaneous O 2 -sensing probes, and fluorine magnetic resonance imaging methods are limited by their intermittent measurements, requirements for skilled personnel, indirect interfaces, and/or their tethered connections. This paper reports a wireless, miniaturized, minimally invasive near-infrared spectroscopic system designed for uninterrupted monitoring of local-tissue oxygenation. A bioresorbable barbed structure anchors the probe stably at implantation sites for a time period matched to the clinical need, with the ability for facile removal afterward. The probe connects to a skin-interfaced electronic module for wireless access to essential physiological parameters, including local tissue oxygenation, pulse oxygenation, and heart rate. In vitro tests and in vivo studies in porcine flap and kidney models demonstrate the ability of the system to continuously measure oxygenation with high accuracy and sensitivity.

Improvements have been made in the diagnosis of Alzheimer’s disease (AD), manifesting mostly in the development of in vivo imaging methods that allow for the detection of pathological changes in AD by magnetic resonance imaging (MRI) and positron emission tomography (PET) scans. Many of these imaging methods, however, use agents that probe amyloid fibrils and plaques–species that do not correlate well with disease progression and are not present at the earliest stages of the disease. Amyloid β oligomers (AβOs), rather, are now widely accepted as the Aβ species most germane to AD onset and progression. Here we report evidence further supporting the role of AβOs as pathological instigators of AD and introduce promising anti-AβO diagnostic probes capable of distinguishing the 5xFAD mouse model from wild type mice by PET and MRI. In a developmental study, Aβ oligomers in 5xFAD mice were found to appear at 3 months of age, just prior to the onset of memory dysfunction, and spread as memory worsened. The increase of AβOs is prominent in the subiculum and correlates with concomitant development of reactive astrocytosis. The impact of these AβOs on memory is in harmony with findings that intraventricular injection of synthetic AβOs into wild type mice induced hippocampal dependent memory dysfunction within 24 h. Compelling support for the conclusion that endogenous AβOs cause memory loss was found in experiments showing that intranasal inoculation of AβO-selective antibodies into 5xFAD mice completely restored memory function, measured 30–40 days post-inoculation. These antibodies, which were modified to give MRI and PET imaging probes, were able to distinguish 5xFAD mice from wild type littermates. These results provide strong support for the role of AβOs in instigating memory loss and salient AD neuropathology, and they demonstrate that AβO selective antibodies have potential both for therapeutics and for diagnostics.

Microelectromechanical systems (MEMS) are essential components in many electronic technologies for consumer and industrial applications. Such devices are typically made using materials selected to support long operational lifetimes, but MEMS designed to physically disintegrate or to dissolve after a targeted period could provide a route to reduce electronic waste and could enable applications that require a finite operating timeframe, such as temporary medical implants. Here we report ecoresorbable and bioresorbable MEMS that are based on fully water-soluble material platforms and can either naturally resorb into the environment to eliminate solid waste or in the body to avoid a need for surgical extraction. We illustrate the biocompatibility of the approach with mechanobiology, histology and haematology studies of the implanted devices and their dissolution end products. We also demonstrate bioresorbable encapsulating materials and deployment strategies in small animal models to reduce device damage, confine mobile fragments and provide robust adhesion with adjacent tissues.

In vivo optogenetics and photopharmacology are two techniques for controlling neuronal activity that have immense potential in neuroscience research. Their applications in tether-free groups of animals have been limited in part due to tools availability. Here, we present a wireless, battery-free, programable multilateral optofluidic platform with user-selected modalities for optogenetics, pharmacology and photopharmacology. This system features mechanically compliant microfluidic and electronic interconnects, capabilities for dynamic control over the rates of drug delivery and real-time programmability, simultaneously for up to 256 separate devices in a single cage environment. Our behavioral experiments demonstrate control of motor behaviors in grouped mice through in vivo optogenetics with co-located gene delivery and controlled photolysis of caged glutamate. These optofluidic systems may expand the scope of wireless techniques to study neural processing in animal models.

Genetic engineering and implantable bioelectronics have transformed investigations of cardiovascular physiology and disease. However, the two approaches have been difficult to combine in the same species: genetic engineering is applied primarily in rodents, and implantable devices generally require larger animal models. We recently developed several miniature cardiac bioelectronic devices suitable for mice and rats to enable the advantages of molecular tools and implantable devices to be combined. Successful implementation of these device-enabled studies requires microsurgery approaches that reliably interface bioelectronics to the beating heart with minimal disruption to native physiology. Here we describe how to perform an open thoracic surgical technique for epicardial implantation of wireless cardiac pacemakers in adult rats that has lower mortality than transvenous implantation approaches. In addition, we provide the methodology for a full biocompatibility assessment of the physiological response to the implanted device. The surgical implantation procedure takes ~40 min for operators experienced in microsurgery to complete, and six to eight surgeries can be completed in 1 d. Implanted pacemakers provide programmed electrical stimulation for over 1 month. This protocol has broad applications to harness implantable bioelectronics to enable fully conscious in vivo studies of cardiovascular physiology in transgenic rodent disease models.