Bioelectronic modulation of carotid sinus nerve to treat type 2 diabetes: current knowledge and future perspectives.

Bioelectronic medicine are an emerging class of treatments aiming to modulate body nervous activity to correct pathological conditions and restore health. Recently, it was shown that the high frequency electrical neuromodulation of the carotid sinus nerve (CSN), a small branch of the glossopharyngeal nerve that connects the carotid body (CB) to the brain, restores metabolic function in type 2 diabetes (T2D) animal models highlighting its potential as a new therapeutic modality to treat metabolic diseases in humans. In this manuscript, we review the current knowledge supporting the use of neuromodulation of the CSN to treat T2D and discuss the future perspectives for its clinical application. Firstly, we review in a concise manner the role of CB chemoreceptors and of CSN in the pathogenesis of metabolic diseases. Secondly, we describe the findings supporting the potential therapeutic use of the neuromodulation of CSN to treat T2D, as well as the feasibility and reversibility of this approach. A third section is devoted to point up the advances in the neural decoding of CSN activity, in particular in metabolic disease states, that will allow the development of closed-loop approaches to deliver personalized and adjustable treatments with minimal side effects. And finally, we discuss the findings supporting the assessment of CB activity in metabolic disease patients to screen the individuals that will benefit therapeutically from this bioelectronic approach in the future.

Bioelectronic Medicine: a multidisciplinary roadmap from biophysics to precision therapies.

Bioelectronic Medicine stands as an emerging field that rapidly evolves and offers distinctive clinical benefits, alongside unique challenges. It consists of the modulation of the nervous system by precise delivery of electrical current for the treatment of clinical conditions, such as post-stroke movement recovery or drug-resistant disorders. The unquestionable clinical impact of Bioelectronic Medicine is underscored by the successful translation to humans in the last decades, and the long list of preclinical studies. Given the emergency of accelerating the progress in new neuromodulation treatments (i.e., drug-resistant hypertension, autoimmune and degenerative diseases), collaboration between multiple fields is imperative. This work intends to foster multidisciplinary work and bring together different fields to provide the fundamental basis underlying Bioelectronic Medicine. In this review we will go from the biophysics of the cell membrane, which we consider the inner core of neuromodulation, to patient care. We will discuss the recently discovered mechanism of neurotransmission switching and how it will impact neuromodulation design, and we will provide an update on neuronal and glial basis in health and disease. The advances in biomedical technology have facilitated the collection of large amounts of data, thereby introducing new challenges in data analysis. We will discuss the current approaches and challenges in high throughput data analysis, encompassing big data, networks, artificial intelligence, and internet of things. Emphasis will be placed on understanding the electrochemical properties of neural interfaces, along with the integration of biocompatible and reliable materials and compliance with biomedical regulations for translational applications. Preclinical validation is foundational to the translational process, and we will discuss the critical aspects of such animal studies. Finally, we will focus on the patient point-of-care and challenges in neuromodulation as the ultimate goal of bioelectronic medicine. This review is a call to scientists from different fields to work together with a common endeavor: accelerate the decoding and modulation of the nervous system in a new era of therapeutic possibilities.

Chronic Intermittent Hypoxia-Induced Dysmetabolism Is Associated with Hepatic Oxidative Stress, Mitochondrial Dysfunction and Inflammation.

The association between obstructive sleep apnea (OSA) and metabolic disorders is well-established; however, the underlying mechanisms that elucidate this relationship remain incompletely understood. Since the liver is a major organ in the maintenance of metabolic homeostasis, we hypothesize that liver dysfunction plays a crucial role in the pathogenesis of metabolic dysfunction associated with obstructive sleep apnea (OSA). Herein, we explored the underlying mechanisms of this association within the liver. Experiments were performed in male Wistar rats fed with a control or high fat (HF) diet (60% lipid-rich) for 12 weeks. Half of the groups were exposed to chronic intermittent hypoxia (CIH) (30 hypoxic (5% O2) cycles, 8 h/day) that mimics OSA, in the last 15 days. Insulin sensitivity and glucose tolerance were assessed. Liver samples were collected for evaluation of lipid deposition, insulin signaling, glucose homeostasis, hypoxia, oxidative stress, antioxidant defenses, mitochondrial biogenesis and inflammation. Both the CIH and HF diet induced dysmetabolism, a state not aggravated in animals submitted to HF plus CIH. CIH aggravates hepatic lipid deposition in obese animals. Hypoxia-inducible factors levels were altered by these stimuli. CIH decreased the levels of oxidative phosphorylation complexes in both groups and the levels of SOD-1. The HF diet reduced mitochondrial density and hepatic antioxidant capacity. The CIH and HF diet produced alterations in cysteine-related thiols and pro-inflammatory markers. The results obtained suggest that hepatic mitochondrial dysfunction and oxidative stress, leading to inflammation, may be significant factors contributing to the development of dysmetabolism associated with OSA.

Effect of Carotid Body Denervation on Systemic Endothelial Function in a Diabetic Animal Model.

Endothelial dysfunction is an essential intermediary for development of cardiovascular diseases associated with diabetes and hypertension (HT). The carotid body (CB) dysfunction contributes to dysmetabolic states, and the resection of carotid sinus nerve (CSN) prevents and reverts dysmetabolism and HT. Herein, we investigated if CSN denervation ameliorates systemic endothelial dysfunction in an animal model of type 2 diabetes mellitus (T2DM).We used Wistar male rats submitted to HFHSu diet during 25 weeks and the correspondent age-matched controls fed with a standard diet. CSN resection was performed in half of the groups after 14 weeks of diet. In vivo insulin sensitivity, glucose tolerance and blood pressure, ex vivo aortic artery contraction and relaxation and nitric oxide (NO) levels in plasma and aorta, aorta nitric oxide synthase (NOS) isoforms, and PGF2αR levels were evaluated.We demonstrated that, alongside to dysmetabolism and HT reversion, CSN resection restores endothelial function in the aorta and decreases the NO levels in plasma and aorta at the same time that restores normal levels of iNOS in aorta without changing eNOS or PGF2αR levels.These results suggest that the modulation of CB activity can be important for the treatment of HT and endothelial dysfunction related with T2DM.

Increased Abdominal Perimeter Differently Affects Respiratory Function in Men and Women.

Obesity is a worldwide epidemic being the main cause of cardiovascular, metabolic disturbances and chronic pulmonary diseases. The increase in body weight may affect the respiratory system due to fat deposition and systemic inflammation. Herein, we evaluated the sex differences in the impact of obesity and high abdominal circumference on basal ventilation. Thirty-five subjects, 23 women and 12 men with a median age of 61 and 67, respectively, were studied and classified as overweight and obese according to body mass index (BMI) and were also divided by the abdominal circumference. Basal ventilation, namely, respiratory frequency, tidal volume, and minute ventilation, was evaluated. In normal and overweight women, basal ventilation did not change, but obese women exhibited a decrease in tidal volume. In men, overweight and obese subjects did not exhibit altered basal ventilation. In contrast, when subjects were subdivided based on the abdominal perimeter, a higher circumference did not change the respiratory frequency but induced a decrease in tidal volume and minute ventilation in women, while in men these two parameters increased. In conclusion, higher abdominal circumference rather than BMI is associated with alterations in basal ventilation in women and men.

Carotid Body Resection Prevents Short-Term Spatial Memory Decline in Prediabetic Rats Without Changing Insulin Signaling in the Hippocampus and Prefrontal Cortex.

Individuals who develop type 2 diabetes (T2D) at an early age are at higher risk of developing neurodegenerative disorders such as Alzheimer's and Parkinson's disease. A shared dysfunctional characteristic between T2D and these neurodegenerative disorders is insulin resistance. Recently, it was shown that prediabetes animals and patients exhibited increased carotid body (CB) activity. Moreover, these organs are deeply involved in the development of metabolic diseases, since upon abolishment of their activity via carotid sinus nerve (CSN) resection, several dysmetabolic features of T2D were reverted. Herein, we investigated if CSN resection may also prevent cognitive impairment associated with brain insulin resistance. We explored a diet-induced prediabetes animal model where Wistar rats are kept in a high fat-high sucrose (HFHSu) diet for 20 weeks. We evaluated CSN resection effects on behavioral parameters and on insulin signaling-related proteins levels, in the prefrontal cortex and the hippocampus. HFHSu animals exhibited impaired short-term memory evaluated by the y-maze test. Remarkably, CSN resection prevented the development of this phenotype. HFHSu diet or CSN resection did not promote significant alterations in insulin signaling-associated proteins levels. Our findings suggest that CBs modulation might have a role in preventing short-term spatial memory deficits associated with peripheral dysmetabolic states.

Chronic Metformin Administration Does Not Alter Carotid Sinus Nerve Activity in Control Rats.

Metformin is a glucose-lowering, insulin-sensitizing drug that is commonly used in the treatment of type 2 diabetes (T2D). In the last decade, the carotid body (CB) has been described as a metabolic sensor implicated in the regulation of glucose homeostasis, being CB dysfunction crucial for the development of metabolic diseases, such as T2D. Knowing that metformin could activate AMP-activated protein kinase (AMPK) and that AMPK has been described to have an important role in CB hypoxic chemotransduction, herein we have investigated the effect of chronic metformin administration on carotid sinus nerve (CSN) chemosensory activity in basal and hypoxic and hypercapnic conditions in control animals. Experiments were performed in male Wistar rats subjected to 3 weeks of metformin (200 mg/kg) administration in the drinking water. The effect of chronic metformin administration was tested in spontaneous and hypoxic (0% and 5% O2) and hypercapnic (10% CO2) evoked CSN chemosensory activity. Metformin administration for 3 weeks did not modify the basal CSN chemosensory activity in control animals. Moreover, the CSN chemosensory response to intense and moderate hypoxia and hypercapnia was not altered by the chronic metformin administration. In conclusion, chronic metformin administration did not modify chemosensory activity in control animals.

On the origins of sleep disordered breathing, cardiorespiratory and metabolic dysfunction: which came first, the chicken or the egg?

Sleep disordered breathing (SDB) is a complex, sex specific and highly heterogeneous group of respiratory disorders. Nevertheless, sleep fragmentation and repeated fluctuations of arterial blood gases for several hours per night are at the core of the problem; together, they impose significant stress to the organism with deleterious consequences on physical and mental health. SDB increases the risk of obesity, diabetes, depression and anxiety disorders; however, the same health issues are risk factors for SDB. So, which came first, the chicken or the egg? What causes the appearance of the first significant apnoeic events during sleep? These are important questions because although moderate to severe SDB affects ∼500 million adults globally, we still have a poor understanding of the origins of the disease, and the main treatments (and animal models) focus on the symptoms rather than the cause. Because obesity, metabolic dysfunction and stress-related neurological disorders generally appear progressively, we discuss how the development of these diseases can lead to specific anatomical and non-anatomical traits of SDB in males and females while considering the impacts of sex steroids. In light of the growing evidence indicating that the carotid bodies are important sensors of key metabolic and endocrine signals associated with stress and dysmetabolism, we propose that these organs play a key role in the process.

Circulating Dopamine Is Regulated by Dietary Glucose and Controls Glucagon-like 1 Peptide Action in White Adipose Tissue.

Dopamine directly acts in the liver and white adipose tissue (WAT) to regulate insulin signaling, glucose uptake, and catabolic activity. Given that dopamine is secreted by the gut and regulates insulin secretion in the pancreas, we aimed to determine its regulation by nutritional cues and its role in regulating glucagon-like peptide 1 (GLP-1) action in WAT. Solutions with different nutrients were administered to Wistar rats and postprandial dopamine levels showed elevations following a mixed meal and glucose intake. In high-fat diet-fed diabetic Goto-Kakizaki rats, sleeve gastrectomy upregulated dopaminergic machinery, showing the role of the gut in dopamine signaling in WAT. Bromocriptine treatment in the same model increased GLP-1R in WAT, showing the role of dopamine in regulating GLP-1R. By contrast, treatment with the GLP-1 receptor agonist Liraglutide had no impact on dopamine receptors. GLP-1 and dopamine crosstalk was shown in rat WAT explants, since dopamine upregulated GLP-1-induced AMPK activity in mesenteric WAT in the presence of the D2R and D3R inhibitor Domperidone. In human WAT, dopamine receptor 1 (D1DR) and GLP-1R expression were correlated. Our results point out a dietary and gut regulation of plasma dopamine, acting in the WAT to regulate GLP-1 action. Together with the known dopamine action in the pancreas, such results may identify new therapeutic opportunities to improve metabolic control in metabolic disorders.

Acute effects of insulin and insulin-induced hypoglycaemia on carotid body chemoreceptor activity and cardiorespiratory responses in dogs.

NEW FINDINGS: What is the central question of this study? What are the effects of insulin and insulin-induced hypoglycaemia on carotid body chemoreceptor activity in vivo and how do carotid body chemoreceptor stimulation-mediated cardiorespiratory responses in beagle dogs compare during euglycaemia and insulin-induced hypoglycaemia? What is the main finding and its importance? Intracarotid insulin administration leads to sustained increase in carotid body chemoreceptor activity and respiratory response with significant cardiovascular effects. Insulin-induced hypoglycaemia exacerbated NaCN-mediated carotid body chemoreceptor activity and respiratory response with enhanced cardiovascular reflex response. These findings suggest that insulin-induced hypoglycaemia augments the carotid body chemoreceptors to initiate the adaptive counter-regulatory responses to restore the normoglycaemic condition. ABSTRACT: The carotid body chemoreceptors (CBC) play an important role in the adaptive counter-regulatory response to hypoglycaemia by evoking the CBC-mediated sympathetic neuronal system to restore normoglycaemia. Ex vivo studies have shown varied responses of insulin-induced hypoglycaemia on CBC function, and several in vivo studies have indirectly established the role of CBCs in restoring normoglycaemia in both animals and humans. However, a direct effect of insulin and/or insulin-induced hypoglycaemia on CBC activity is not established in animal models. Therefore, the aim of this study was to evaluate in vivo effects of insulin and insulin-induced hypoglycaemia on CBC activity and cardiorespiration in a preclinical large animal model. The carotid sinus nerve (CSN) activity and cardiorespiratory responses to sodium cyanide (NaCN; 25 µg/kg) were compared before (euglycaemic) and after (hypoglycaemic) intracarotid administration of insulin (12.5-100 µU/dogs) in beagle dogs. Insulin administration increased CSN activity and minute ventilation ( V ̇ $\dot V$ E ) with significant (P < 0.0001) effects on heart rate and blood pressure. Insulin-mediated effects on CSN and cardiorespiration were sustained and the change in V ̇ $\dot V$ E was driven by tidal volume only. Insulin significantly (P < 0.0001) lowered blood glucose level. NaCN-mediated CSN activity and V ̇ $\dot V$ E were significantly (P < 0.0001) augmented during insulin-induced hypoglycaemia. The augmented V ̇ $\dot V$ E was primarily driven by respiratory frequency and partially by tidal volume. The cardiovascular reflex response mediated through CBC stimulation was significantly (P < 0.0001) exacerbated during insulin-induced hypoglycaemia. Collectively, these results demonstrate direct effects of insulin and insulin-induced hypoglycaemia on CBC chemosensitivity to potentiate CBC-mediated neuroregulatory pathways to initiate adaptive neuroendocrine and cardiorespiratory counter-regulatory responses to restore normoglycaemia.

Overnutrition during Pregnancy and Lactation Induces Gender-Dependent Dysmetabolism in the Offspring Accompanied by Heightened Stress and Anxiety.

Maternal obesity and gestational diabetes predispose the next generation to metabolic disturbances. Moreover, the lactation phase also stands as a critical phase for metabolic programming. Nevertheless, the precise mechanisms originating these changes remain unclear. Here, we investigate the consequences of a maternal lipid-rich diet during gestation and lactation and its impact on metabolism and behavior in the offspring. Two experimental groups of Wistar female rats were used: a control group (NC) that was fed a standard diet during the gestation and lactation periods and an overnutrition group that was fed a high-fat diet (HF, 60% lipid-rich) during the same phases. The offspring were analyzed at postnatal days 21 and 28 and at 2 months old (PD21, PD28, and PD60) for their metabolic profiles (weight, fasting glycemia insulin sensitivity, and glucose tolerance) and euthanized for brain collection to evaluate metabolism and inflammation in the hypothalamus, hippocampus, and prefrontal cortex using Western blot markers of synaptic dynamics. At 2 months old, behavioral tests for anxiety, stress, cognition, and food habits were conducted. We observed that the female offspring born from HF mothers exhibited increased weight gain and decreased glucose tolerance that attenuated with age. In the offspring males, weight gain increased at P21 and worsened with age, while glucose tolerance remained unchanged. The offspring of the HF mothers exhibited elevated levels of anxiety and stress during behavioral tests, displaying decreased predisposition for curiosity compared to the NC group. In addition, the offspring from mothers with HF showed increased food consumption and a lower tendency towards food-related aggression. We conclude that exposure to an HF diet during pregnancy and lactation induces dysmetabolism in the offspring and is accompanied by heightened stress and anxiety. There was sexual dimorphism in the metabolic traits but not behavioral phenotypes.

A Metabolic Signature to Monitor Endothelial Cell Differentiation, Activation, and Vascular Organization.

The formation of new blood vessels is an important step in the morphogenesis and organization of tissues and organs; hence, the success of regenerative medicine procedures is highly dependent on angiogenesis control. Despite the biotechnological advances, tissue engineering is still a challenge. Regarding vascular network formation, the regulators are well known, yet the identification of markers is pivotal in order to improve the monitoring of the differentiation and proliferation of endothelial cells, as well as the establishment of a vascular network supporting tissue viability for an efficacious implantation. The metabolic profile accompanies the physiological stages of cells involved in angiogenesis, being a fruitful hub of biomarkers, whose levels can be easily retrieved. Through NMR spectroscopy, we identified branched amino acids, acetate, and formate as central biomarkers of monocyte-to-endothelial-cell differentiation and endothelial cell proliferation. This study reinforces the successful differentiation process of monocytes into endothelial cells, allowing self-to-self transplantation of patient-derived vascular networks, which is an important step in tissue engineering, since monocytes are easily isolated and autologous transplantation reduces the immune rejection events.

Impact of Diet Composition on Insulin Resistance.

Insulin resistance is a complex condition in which the body does not respond adequately to insulin, a hormone secreted by the pancreas with an essential role in the regulation of blood sugar levels [...].

Hypoxic and Hypercapnic Responses in Transgenic Murine Model of Alzheimer's Disease Overexpressing Human AßPP: The Effects of Pretreatment with Memantine and Rivastigmine.

Despite the severe respiratory problems reducing the quality of life for Alzheimer's disease (AD) patients, their causes are poorly understood. We aimed to investigate hypoxic and hypercapnic respiratory responses in a transgenic mouse model of AD (AßPP V717I) overexpressing AßPP and mimicking early-onset AD. The cholinesterase inhibitor rivastigmine and the NMDA receptor antagonist memantine were used to investigate the effects of drugs, used to treat AD cognitive dysfunction, on breathing in hypoxia and hypercapnia. We found a significant increase in the respiratory response to hypercapnia and no difference in the hypoxic response in APP+ mice, compared with the control group (APP-). Memantine had no effect on respiration in either group, including responses to hypoxia and hypercapnia. Rivastigmine depressed resting ventilation and response to hypercapnia irrespective of the mice genotype. Reduction in hypoxia-augmented ventilation by rivastigmine was observed only in APP+ mice, which exhibited lower acetylcholinesterase activity in the hippocampus. Treatment with rivastigmine reduced the enzyme activity in both groups equally in the hippocampus and brainstem. The increased ventilatory response to hypercapnia in transgenic mice may indicate alterations in chemoreceptive respiratory nuclei, resulting in increased CO2 sensitivity. Rivastigmine is a potent reductant of normoxic and hypercapnic respiration in APP+ and APP- mice.

Long-Term Hypercaloric Diet Consumption Exacerbates Age-Induced Dysmetabolism and Carotid Body Dysfunction: Beneficial Effects of CSN Denervation.

Carotid bodies (CBs) are metabolic sensors whose dysfunction is involved in the genesis of dysmetabolic states. Ageing induces significant alterations in CB function also prompting to metabolic deregulation. On the other hand, metabolic disease can accelerate ageing processes. Taking these into account, we evaluated the effect of long-term hypercaloric diet intake and CSN resection on age-induced dysmetabolism and CB function. Experiments were performed in male Wistar rats subjected to 14 or 44 weeks of high-fat high-sucrose (HFHSu) or normal chow (NC) diet and subjected to either carotid sinus nerve (CSN) resection or a sham procedure. After surgery, the animals were kept on a diet for more than 9 weeks. Metabolic parameters, basal ventilation, and hypoxic and hypercapnic ventilatory responses were evaluated. CB type I and type II cells, HIF-1α and insulin receptor (IR), and GLP-1 receptor (GLP1-R)-positive staining were analyzed by immunofluorescence. Ageing decreased by 61% insulin sensitivity in NC animals, without altering glucose tolerance. Short-term and long-term HFHSu intake decreased insulin sensitivity by 55 and 62% and glucose tolerance by 8 and 29%, respectively. CSN resection restored insulin sensitivity and glucose tolerance. Ageing decreased spontaneous ventilation, but short-term or long-term intake of HFHSu diet and CSN resection did not modify basal ventilatory parameters. HFHSu diet increased hypoxic ventilatory responses in young and adult animals, effects attenuated by CSN resection. Ageing, hypercaloric diet, and CSN resection did not change hypercapnic ventilatory responses. Adult animals showed decreased type I cells and IR and GLP-1R staining without altering the number of type II cells and HIF-1α. HFHSu diet increased the number of type I and II cells and IR in young animals without significantly changing these values in adult animals. CSN resection restored the number of type I cells in HFHSu animals and decreased IR-positive staining in all the groups of animals, without altering type II cells, HIF-1α, or GLP-1R staining. In conclusion, long-term hypercaloric diet consumption exacerbates age-induced dysmetabolism, and both short- and long-term hypercaloric diet intakes promote significant alterations in CB function. CSN resection ameliorates these effects. We suggest that modulation of CB activity is beneficial in exacerbated stages of dysmetabolism.

Dysmetabolism and Sleep Fragmentation in Obstructive Sleep Apnea Patients Run Independently of High Caffeine Consumption.

Daytime hypersomnolence, the prime feature of obstructive sleep apnea (OSA), frequently leads to high coffee consumption. Nevertheless, some clinicians ask for patients' caffeine avoidance. Caffeinated drinks are sometimes associated with more severe OSA. However, these effects are not consensual. Here we investigated the effect of caffeine consumption on sleep architecture and apnea/hypopnea index in OSA. Also, the impact of caffeine on variables related with dysmetabolism, dyslipidemia, and sympathetic nervous system (SNS) dysfunction were investigated. A total of 65 patients diagnosed with OSA and 32 without OSA were included after given written informed consent. Polysomnographic studies were performed. Blood was collected to quantify caffeine and its metabolites in plasma and biochemical parameters. 24 h urine samples were collected for catecholamines measurement. Statistical analyses were performed by SPSS: (1) non-parametric Mann-Whitney test to compare variables between controls and OSA; (2) multivariate logistic regression testing the effect of caffeine on sets of variables in the 2 groups; and (3) Spearmans' correlation between caffeine levels and comorbidities in patients with OSA. As expected OSA development is associated with dyslipidemia, dysmetabolism, SNS dysfunction, and sleep fragmentation. There was also a significant increase in plasma caffeine levels in the OSA group. However, the higher consumption of caffeine by OSA patients do not alter any of these associations. These results showed that there is no apparent rationale for caffeine avoidance in chronic consumers with OSA.

Dysmetabolism and Neurodegeneration: Trick or Treat?

Accumulating evidence suggests the existence of a strong link between metabolic syndrome and neurodegeneration. Indeed, epidemiologic studies have described solid associations between metabolic syndrome and neurodegeneration, whereas animal models contributed for the clarification of the mechanistic underlying the complex relationships between these conditions, having the development of an insulin resistance state a pivotal role in this relationship. Herein, we review in a concise manner the association between metabolic syndrome and neurodegeneration. We start by providing concepts regarding the role of insulin and insulin signaling pathways as well as the pathophysiological mechanisms that are in the genesis of metabolic diseases. Then, we focus on the role of insulin in the brain, with special attention to its function in the regulation of brain glucose metabolism, feeding, and cognition. Moreover, we extensively report on the association between neurodegeneration and metabolic diseases, with a particular emphasis on the evidence observed in animal models of dysmetabolism induced by hypercaloric diets. We also debate on strategies to prevent and/or delay neurodegeneration through the normalization of whole-body glucose homeostasis, particularly via the modulation of the carotid bodies, organs known to be key in connecting the periphery with the brain.