Biotransformation research advances - 2022 year in review.

This annual review is the eighth of its kind since 2016 (Baillie et al. 2016, Khojasteh et al. 2017, Khojasteh et al. 2018, Khojasteh et al. 2019, Khojasteh et al. 2020, Khojasteh et al. 2021, Khojasteh et al. 2022). Our objective is to explore and share articles which we deem influential and significant in the field of biotransformation.

Bioactivation and reactivity research advances - 2022 year in review‡.

With the 50th year mark since the launch of Drug Metabolism and Disposition journal, the field of drug metabolism and bioactivation has advanced exponentially in the past decades (Guengerich 2023).This has, in a major part, been due to the continued advances across the whole spectrum of applied technologies in hardware, software, machine learning (ML), and artificial intelligence (AI). LC-MS platforms continue to evolve to support key applications in the field, and automation is also improving the accuracy, precision, and throughput of these supporting assays. In addition, sample generation and processing is being aided by increased diversity and quality of reagents and bio-matrices so that what is being analyzed is more relevant and translatable. The application of in silico platforms (applied software, ML, and AI) is also making great strides, and in tandem with the more traditional approaches mentioned previously, is significantly advancing our understanding of bioactivation pathways and how these play a role in toxicity. All of this continues to allow the area of bioactivation to evolve in parallel with associated fields to help bring novel or improved medicines to patients with urgent or unmet needs.Shuai Wang and Cyrus Khojasteh, on behalf of the authors.

The Absolute Bioavailability and Absorption, Metabolism, and Excretion of Ipatasertib, a Potent and Highly Selective Protein Kinase B (Akt) Inhibitor.

Ipatasertib (GDC-0068) is a potent, highly selective, small-molecule inhibitor of protein kinase B (Akt) being developed by Genentech/Roche as a single agent and in combination with other therapies for the treatment of cancers. To fully understand the absorption, metabolism, and excretion of ipatasertib in humans, an open-label study using 14C-radiolabeled ipatasertib was completed to characterize the absolute bioavailability (period 1) and mass balance and metabolite profiling (period 2). In period 1, subjects were administered a 200 mg oral dose of ipatasertib followed by an 80 µg (800 nCi) intravenous dose of [14C]-ipatasertib. In period 2, subjects received a single oral dose containing approximately 200 mg (100 µCi) [14C]-ipatasertib. In an integrated analytical strategy, accelerator mass spectrometry was applied to measure the 14C microtracer intravenous pharmacokinetics in period 1 and fully profile plasma radioactivity in period 2. The systemic plasma clearance and steady-state volume of distribution were 98.8 L/h and 2530 L, respectively. The terminal half-lives after oral and intravenous administrations were similar (26.7 and 27.4 hours, respectively) and absolute bioavailability of ipatasertib was 34.0%. After a single oral dose of [14C]-ipatasertib, 88.3% of the administered radioactivity was recovered with approximately 69.0% and 19.3% in feces and urine, respectively. Radioactivity in feces and urine was predominantly metabolites with 24.4% and 8.26% of dose as unchanged parent, respectively; indicating that ipatasertib had been extensively absorbed and hepatic metabolism was the major route of clearance. The major metabolic pathway was N-dealkylation mediated by CYP3A, and minor pathways were oxidative by cytochromes P450 and aldehyde oxidase. SIGNIFICANCE STATEMENT: The study provided definitive information regarding the absolute bioavailability and the absorption, metabolism, and excretion pathways of ipatasertib, a potent, novel, and highly selective small-molecule inhibitor of protein kinase B (Akt). An ultrasensitive radioactive counting method, accelerator mass spectrometry was successfully applied for 14C-microtracer absolute bioavailability determination and plasma metabolite profiling.

Phenylpropionic acid produced by gut microbiota alleviates acetaminophen-induced hepatotoxicity.

The gut microbiota affects hepatic drug metabolism. However, gut microbial factors modulating hepatic drug metabolism are largely unknown. In this study, using a mouse model of acetaminophen (APAP)-induced hepatotoxicity, we identified a gut bacterial metabolite that controls the hepatic expression of CYP2E1 that catalyzes the conversion of APAP to a reactive, toxic metabolite. By comparing C57BL/6 substrain mice from two different vendors, Jackson (6J) and Taconic (6N), which are genetically similar but harbor different gut microbiotas, we established that the differences in the gut microbiotas result in differential susceptibility to APAP-induced hepatotoxicity. 6J mice exhibited lower susceptibility to APAP-induced hepatotoxicity than 6N mice, and such phenotypic difference was recapitulated in germ-free mice by microbiota transplantation. Comparative untargeted metabolomic analysis of portal vein sera and liver tissues between conventional and conventionalized 6J and 6N mice led to the identification of phenylpropionic acid (PPA), the levels of which were higher in 6J mice. PPA supplementation alleviated APAP-induced hepatotoxicity in 6N mice by lowering hepatic CYP2E1 levels. Moreover, PPA supplementation also reduced carbon tetrachloride-induced liver injury mediated by CYP2E1. Our data showed that previously known PPA biosynthetic pathway is responsible for PPA production. Surprisingly, while PPA in 6N mouse cecum contents is almost undetectable, 6N cecal microbiota produces PPA as well as 6J cecal microbiota in vitro, suggesting that PPA production in the 6N gut microbiota is suppressed in vivo. However, previously known gut bacteria harboring the PPA biosynthetic pathway were not detected in either 6J or 6N microbiota, suggesting the presence of as-yet-unidentified PPA-producing gut microbes. Collectively, our study reveals a novel biological function of the gut bacterial metabolite PPA in the gut-liver axis and presents a critical basis for investigating PPA as a modulator of CYP2E1-mediated liver injury and metabolic diseases.

Dissecting the human gut microbiome to better decipher drug liability: A once-forgotten organ takes center stage.

BACKGROUND: The gut microbiome is a diverse system within the gastrointestinal tract composed of trillions of microorganisms (gut microbiota), along with their genomes. Accumulated evidence has revealed the significance of the gut microbiome in human health and disease. Due to its ability to alter drug/xenobiotic pharmacokinetics and therapeutic outcomes, this once-forgotten "metabolic organ" is receiving increasing attention. In parallel with the growing microbiome-driven studies, traditional analytical techniques and technologies have also evolved, allowing researchers to gain a deeper understanding of the functional and mechanistic effects of gut microbiome. AIM OF REVIEW: From a drug development perspective, microbial drug metabolism is becoming increasingly critical as new modalities (e.g., degradation peptides) with potential microbial metabolism implications emerge. The pharmaceutical industry thus has a pressing need to stay up-to-date with, and continue pursuing, research efforts investigating clinical impact of the gut microbiome on drug actions whilst integrating advances in analytical technology and gut microbiome models. Our review aims to practically address this need by comprehensively introducing the latest innovations in microbial drug metabolism research- including strengths and limitations, to aid in mechanistically dissecting the impact of the gut microbiome on drug metabolism and therapeutic impact, and to develop informed strategies to address microbiome-related drug liability and minimize clinical risk. KEY SCIENTIFIC CONCEPTS OF REVIEW: We present comprehensive mechanisms and co-contributing factors by which the gut microbiome influences drug therapeutic outcomes. We highlight in vitro, in vivo, and in silico models for elucidating the mechanistic role and clinical impact of the gut microbiome on drugs in combination with high-throughput, functionally oriented, and physiologically relevant techniques. Integrating pharmaceutical knowledge and insight, we provide practical suggestions to pharmaceutical scientists for when, why, how, and what is next in microbial studies for improved drug efficacy and safety, and ultimately, support precision medicine formulation for personalized and efficacious therapies.

Strain-Driven Negative Resistance Switching of Conductive Fibers with Adjustable Sensitivity for Wearable Healthcare Monitoring Systems with Near-Zero Standby Power.

Recently, one of the primary concerns in e-textile-based healthcare monitoring systems for chronic illness patients has been reducing wasted power consumption, as the system should be always-on to capture diverse biochemical and physiological characteristics. However, the general conductive fibers, a major component of the existing wearable monitoring systems, have a positive gauge-factor (GF) that increases electrical resistance when stretched, so that the systems have no choice but to consume power continuously. Herein, a twisted conductive-fiber-based negatively responsive switch-type (NRS) strain-sensor with an extremely high negative GF (resistance change ratio ≈ 3.9 × 108 ) that can significantly increase its conductivity from insulating to conducting properties is developed. To this end, a precision cracking technology is devised, which could induce a difference in the Young's modulus of the encapsulated layer on the fiber through selective ultraviolet-irradiation treatment. Owing to this technology, the NRS strain-sensors can allow for effective regulation of the mutual contact resistance under tensile strain while maintaining superior durability for over 5000 stretching cycles. For further practical demonstrations, three healthcare monitoring systems (E-fitness pants, smart-masks, and posture correction T-shirts) with near-zero standby power are also developed, which opens up advancements in electronic textiles by expanding the utilization range of fiber strain-sensors.

Strain-Insensitive Stretchable Fiber Conductors Based on Highly Conductive Buckled Shells for Wearable Electronics.

Based on their high applicability to wearable electronics, fiber-based stretchable electronics have been developed via different strategies. However, the electrical conductivity of a fiber electrode is severely degraded, following deformation upon stretching. Despite the introduction of conductive buckled structures to resolve this issue, there still exist limitations regarding the simultaneous realizations of high conductivity and stretchability. Here, we exploit the dense distribution of the Ag nanoparticle (AgNP) network in polyurethane (PU) to fabricate a strain-insensitive stretchable fiber conductor comprising highly conductive buckled shells via a facile chemical process. These buckled AgNPs/PU fibers exhibit stable and reliable electrical responses across a wide range (tensile strain = ∼200%), in addition to their high electrical conductivity (26,128 S/m) and quality factor (Q = 2.29). Particularly, the negligible electrical hysteresis and excellent durability (>10,000 stretching-releasing cycles) of the fibers demonstrate their high applicability to wearable electronics. Furthermore, we develop buckled fiber-based pH sensors exhibiting stable, repeatable, and highly distinguishable responses (changing pH is from 4 to 8, response time is 5-6 s) even under 100% tensile strain. The buckled AgNPs/PU fibers represent a facile strategy for maintaining the stable electrical performances of fiber electrodes across the strain range of human motion for wearable applications.

Absorption, Metabolism, and Excretion of Taselisib (GDC-0032), a Potent ß-Sparing PI3K Inhibitor in Rats, Dogs, and Humans.

Taselisib (also known as GDC-0032) is a potent and selective phosphoinositide 3-kinase (PI3K) inhibitor that displays greater selectivity for mutant PI3Kα than wild-type PI3Kα To better understand the absorption, distribution, metabolism, and excretion properties of taselisib, mass balance studies were conducted following single oral doses of [14C]taselisib in rats, dogs, and humans. Absolute bioavailability (ABA) of taselisib in humans was determined by oral administration of taselisib at the therapeutic dose followed by intravenous dosing of [14C]taselisib as a microtracer. The ABA in humans was 57.4%. Absorption of taselisib was rapid in rats and dogs and moderately slow in humans. The recovery of radioactivity in excreta was high (>96%) in the three species where feces was the major route of excretion. Taselisib was the major circulating component in the three species with no metabolite accounting for >10% of the total drug-derived material. The fraction absorbed of taselisib was 35.9% in rats and 71.4% in dogs. In rats, absorbed drug underwent moderate to extensive metabolism and biliary excretion of taselisib was minor. In dog, biliary excretion and metabolism were major clearance pathways. In humans, 84.2% of the dose was recovered as the parent drug in excreta indicating that metabolism played a minor role in the drug's clearance. Major metabolism pathways were oxidation and amide hydrolysis in the three species while methylation was another prominent metabolism pathway in dogs. The site of methylation was identified on the triazole moiety. In vitro experiments characterized that the N-methylation was dog-specific and likely mediated by a thiol methyltransferase. SIGNIFICANCE STATEMENT: This study provides a comprehensive description of the absorption, distribution, and metabolism and pharmacokinetic properties of taselisib in preclinical species and humans. This study demonstrated the importance of oral bioavailability results for understanding taselisib's clearance pathways. The study also describes the identification and characterization of a unique dog-specific N-methylation metabolite of taselisib and the enzyme mediating N-methylation in vitro.

Biotransformation novel advances - 2021 year in review.

Biotransformation field is constantly evolving with new molecular structures and discoveries of metabolic pathways that impact efficacy and safety. Recent review by Kramlinger et al. (2022) nicely captures the future (and the past) of highly impactful science of biotransformation (see the first article). Based on the selected articles, this review was categorized into three sections: (1) new modalities biotransformation, (2) drug discovery biotransformation, and (3) drug development biotransformation (Table 1).

Bioactivation and reactivity research advances - 2021 year in review.

This year's review on bioactivation and reactivity began as a part of the annual review on biotransformation and bioactivation led by Cyrus Khojasteh (see references). Increased contributions from experts in the field led to the development of a stand alone edition for the first time this year focused specifically on bioactivation and reactivity. Our objective for this review is to highlight and share articles which we deem influential and significant regarding the development of covalent inhibitors, mechanisms of reactive metabolite formation, enzyme inactivation, and drug safety. Based on the selected articles, we created two sections: (1) reactivity and enzyme inactivation, and (2) bioactivation mechanisms and safety (Table 1). Several biotransformation experts have contributed to this effort from academic and industry settings.[Table: see text].

Elucidating the structure and cytochrome P450-mediated mechanism for novel metabolites of GDC-0575 in rats.

1. GDC-0575 is an ATP-competitive small-molecule inhibitor of ChK1 that is being developed by Genentech for the treatment of various human malignancies.2. In a radiolabeled mass balance study of GDC-0575 in rats, two novel metabolites, named M12 (-71 Da,) and M17 (+288 Da), were detected as abundant circulating metabolites.3. Subsequent mass spectrometry and nuclear magnetic resonance analysis showed that M12 was a cyclized metabolite of GDC-0575, whereas M17 was its heterodimer to the parent. We further determined that M12 was mainly generated by cytochrome P450 (Cyp) 2d2.4. We proposed the potential mechanism was initiated by the oxidation on the pyrrole ring and subsequent cyclisation of the free primary amine onto C-3 of the pyrrole ring. This was followed by expulsion of cyclopropylcarboxamide and a loss of water to form intermediate I, which can be further oxidised to form M12, or dimerise with another molecule of GDC-0575 as nucleophile to form M17.5. To verify this hypothesis, we attempted to trap the intermediate I with glutathione (GSH) trapping assay and the GSH conjugate on the pyrrole ring was identified. This suggests the oxidation on the pyrrole led to reactive metabolite formation and supported this proposed mechanism.

Post-Traumatic Epilepsy in Zebrafish Is Drug-Resistant and Impairs Cognitive Function.

Post-traumatic epilepsy (PTE) is acquired epilepsy after traumatic brain injury (TBI). Despite the availability of more than 20 antiseizure medications (ASMs), there is no way at present to prevent epileptogenesis in TBI survivors, and many cases of PTE become drug-resistant. Importantly, the adverse effects of ASMs can significantly affect patients' quality of life. Mammalian models are commonly used for studying refractory PTE, but are expensive and laborious. Zebrafish models have become popular for studying epilepsy, but most focus on larvae, and there have been no reports to date of pharmacological screening in an adult zebrafish model of acquired epilepsy. Valid animal models are critical for understanding PTE and for developing novel therapeutics. The aim of the present study was to characterize the cognitive impairments of a zebrafish model of TBI that leads to the development of PTE. Using combined behavioral and electrophysiological approaches, we also characterized the pharmacological effects of the most commonly used ASMs to manage PTE (valproate, carbamazepine, and phenytoin). Zebrafish with PTE exhibited impairments in learning and memory, difficulty in decision making, and reduced social preference. Valproate and carbamazepine had a limited protective effect against behavioral seizures, and all three drugs failed to significantly reduce electrographical seizures. The negative impacts of TBI and ASMs in zebrafish parallel those observed in other animals, making the zebrafish model of PTE a promising high-throughput model of refractory and drug-resistant epilepsy.

Absorption, metabolism and excretion of pictilisib, a potent pan-class I phosphatidylinositol-3-Kinase (PI3K) inhibitor, in rats, dogs, and humans.

The absorption, metabolism and excretion of pictilisib, a selective small molecule inhibitor of class 1 A phosphoinositide 3-kinase (PI3K), was characterized following a single oral administration of [14C]pictilisib in rats, dogs and humans at the target doses of 30 mg/kg, 5 mg/kg and 60 mg, respectively.Pictilisib was rapidly absorbed with Tmax less than 2 h across species. In systemic circulation, pictilisib represented the predominant total radioactivity greater than 86.6% in all species.Total pictilisib and related radioactivity was recovered from urine and faeces in rats, dogs, and human at 98%, 80% and 95%, respectively, with less than 2% excreted in urine and the rest excreted into faeces.In rat and dog, more than 40% of drug-related radioactivity was excreted into the bile suggesting biliary excretion was the major route of excretion. Unchanged pictilisib was a minor component in rat and dog bile. The major metabolite in bile was O-glucuronide of oxidation on indazole moiety (M20, 21% of the dose) in rats and an oxidative piperazinyl ring-opened metabolite M7 (10.8% of the dose) in dogs.Oxidative glutathione (GSH) conjugates (M18, M19) were novel metabolites detected in rat bile, suggesting the potential generation of reactive intermediates from pictilisib. The structure of M18 was further confirmed by NMR to be a N-hydroxylated and GSH conjugated metabolite on the moiety of the indazole ring.

Age Bias in Zebrafish Models of Epilepsy: What Can We Learn From Old Fish?

Zebrafish are a powerful tool for investigating epilepsy. Mammalian seizures can be recapitulated molecularly, behaviorally, and electrophysiologically, using a fraction of the resources required for experiments in mammals. Larval zebrafish offer exceptionally economical and high-throughput approaches and are amenable to state-of-the-art genetic engineering techniques, providing valuable transgenic models of human diseases. For these reasons, larvae tend to be chosen for studying epilepsy, but the value of adult zebrafish may be underappreciated. Zebrafish exhibit transient larval - adult duality. The incompletely developed neural system of larval zebrafish may limit the translation of complex neurological disorders. Larval zebrafish go through dynamic changes during ontogenesis, whereas adult zebrafish are physiologically more stable. Adult zebrafish have a full range of complex brain structures and functions, such as an endothelial blood-brain barrier and adult neurogenesis, both are significant factors in epilepsy research. This review highlights the differences between larval and adult zebrafish that should be considered in pathophysiological and pharmacological studies of epilepsy.

Zebrafish model of posttraumatic epilepsy.

OBJECTIVE: Posttraumatic epilepsy (PTE) is defined as recurrent and unprovoked seizures occurring >1 week after traumatic brain injury (TBI). Animal studies of PTE are lengthy and expensive. In this study, we developed a cost-effective PTE animal model using zebrafish to bridge the gap between in vitro studies and low-throughput animal studies. METHODS: We used two different sets of parameters (G1 and G2) to induce closed-head TBI in adult zebrafish using pulsed high-intensity focused ultrasound. Injured fish and naive controls were evaluated for behavioral deficits and spontaneous behavioral seizure activity up to 21 days postinjury (DPI). We also assessed behavioral seizure susceptibility to a subconvulsive dose of pentylenetetrazole (PTZ; 2.5 mmol·L-1 ) and recorded electrophysiological signals to confirm seizure activity up to 40 DPI. In addition, we investigated injury-related changes in the blood-brain barrier and expression levels of various proteins altered in rodent and human TBI. RESULTS: The G2 parameters resulted in a more severe TBI, with a mortality rate of 25%, as well as motor dysfunction and heightened anxiety persisting at 21 DPI. One hundred percent of the G2 group showed spontaneous myocloniclike behavior, and 80% demonstrated tonic-clonic-like behavioral seizures by 21 DPI. Such activities were not detected in the naive group. After the application of 2.5 mmol·L-1 PTZ, 100% of injured zebrafish had cloniclike seizures at 21 DPI, versus 30% of the naive group. We also demonstrated electrographic seizure activity at 40 DPI, which was not detected in the naive controls. Lastly, we observed acute blood-brain barrier dysfunction and increased levels of HMGB1 and ratios of phosphorylated/total Akt and tau through 21 DPI. SIGNIFICANCE: Together, the results indicate that severe TBI in the adult zebrafish leads to similar behavioral and physiological changes to those of more traditional models, including the development of PTE, and suggest this may be a useful model that can accelerate research in TBI/PTE.

Poor Work-Life Balance May Lead to Impaired Cognitive Function in Bus Drivers.

OBJECTIVE: This study aimed to investigate how work-life balance (WLB) corresponds to cognitive functions and which mental health conditions play a mediating role in this association among Korean bus drivers. METHODS: The cognitive failures questionnaire (CFQ) was administered to 347 bus drivers in Seoul, Korea. The differences in the CFQ and WLB scores were examined by analysis of covariance, and a structural equation model (SEM) was constructed for investigating the mediating role of mental health indices between WLB and CFQ scores. RESULTS: Compared with the highest subjective work-life balance group, the lowest group had significantly higher CFQ scores. In the SEM, anxiety was a mediating variable between subjective work-life balance and CFQ scores. CONCLUSIONS: Work-life balance is associated with cognitive failures among Korean bus drivers, and anxiety was a key mediating mental health indicator.

A 3D-Printed Sensor for Monitoring Biosignals in Small Animals.

Although additive manufacturing technologies, also known as 3D printing, were first introduced in the 1980s, they have recently gained remarkable popularity owing to decreased costs. 3D printing has already emerged as a viable technology in many industries; in particular, it is a good replacement for microfabrication technology. Microfabrication technology usually requires expensive clean room equipment and skilled engineers; however, 3D printing can reduce both cost and time dramatically. Although 3D printing technology has started to emerge into microfabrication manufacturing and medical applications, it is typically limited to creating mechanical structures such as hip prosthesis or dental implants. There have been increased interests in wearable devices and the critical part of such wearable devices is the sensing part to detect biosignals noninvasively. In this paper, we have built a 3D-printed sensor that can measure electroencephalogram and electrocardiogram from zebrafish. Despite measuring biosignals noninvasively from zebrafish has been known to be difficult due to that it is an underwater creature, we were able to successfully obtain electrophysiological information using the 3D-printed sensor. This 3D printing technique can accelerate the development of simple noninvasive sensors using affordable equipment and provide an economical solution to physiologists who are unfamiliar with complicated microfabrication techniques.

Author Correction: Zebrafish as an animal model in epilepsy studies with multichannel EEG recordings.

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

Fructose diet alleviates acetaminophen-induced hepatotoxicity in mice.

Acetaminophen (APAP) is a commonly used analgesic and antipyretic that can cause hepatotoxicity due to production of toxic metabolites via cytochrome P450 (Cyp) 1a2 and Cyp2e1. Previous studies have shown conflicting effects of fructose (the major component in Western diet) on the susceptibility to APAP-induced hepatotoxicity. To evaluate the role of fructose-supplemented diet in modulating the extent of APAP-induced liver injury, male C57BL/6J mice were given 30% (w/v) fructose in water (or regular water) for 8 weeks, followed by oral administration of APAP. APAP-induced liver injury (determined by serum levels of liver enzymes) was decreased by two-fold in mice pretreated with fructose. Fructose-treated mice exhibited (~1.5 fold) higher basal glutathione levels and (~2 fold) lower basal (mRNA and activity) levels of Cyp1a2 and Cyp2e1, suggesting decreased bioactivation of APAP and increased detoxification of toxic metabolite in fructose-fed mice. Hepatic mRNA expression of heat shock protein 70 was also found increased in fructose-fed mice. Analysis of bacterial 16S rRNA gene amplicons from the cecal samples of vehicle groups showed that the fructose diet altered gut bacterial community, leading to increased α-diversity. The abundance of several bacterial taxa including the genus Anaerostipes was found to be significantly correlated with the levels of hepatic Cyp2e1, Cyp1a2 mRNA, and glutathione. Together, these results suggest that the fructose-supplemented diet decreases APAP-induced liver injury in mice, in part by reducing metabolic activation of APAP and inducing detoxification of toxic metabolites, potentially through altered composition of gut microbiota.

Recording nerve signals in canine sciatic nerves with a flexible penetrating microelectrode array.

OBJECTIVE: Previously, we presented the fabrication and characterization of a flexible penetrating microelectrode array (FPMA) as a neural interface device. In the present study, we aim to prove the feasibility of the developed FPMA as a chronic intrafascicular recording tool for peripheral applications. APPROACH: For recording from the peripheral nerves of medium-sized animals, the FPMA was integrated with an interconnection cable and other parts that were designed to fit canine sciatic nerves. The uniformity of tip exposure and in vitro electrochemical properties of the electrodes were characterized. The capability of the device to acquire in vivo electrophysiological signals was evaluated by implanting the FPMA assembly in canine sciatic nerves acutely as well as chronically for 4 weeks. We also examined the histology of implanted tissues to evaluate the damage caused by the device. MAIN RESULTS: Throughout recording sessions, we observed successful multi-channel recordings (up to 73% of viable electrode channels) of evoked afferent and spontaneous nerve unit spikes with high signal quality (SNR > 4.9). Also, minor influences of the device implantation on the morphology of nerve tissues were found. SIGNIFICANCE: The presented results demonstrate the viability of the developed FPMA device in the peripheral nerves of medium-sized animals, thereby bringing us a step closer to human applications. Furthermore, the obtained data provide a driving force toward a further study for device improvements to be used as a bidirectional neural interface in humans.