Physiological Temperature Changes Fine-Tune ß 2- Adrenergic Receptor-Induced Cytosolic cAMP Accumulation.

Norepinephrine (NE) controls many vital body functions by activating adrenergic receptors (ARs). Average core body temperature (CBT) in mice is 37°C. Of note, CBT fluctuates between 36 and 38°C within 24 hours, but little is known about the effects of CBT changes on the pharmacodynamics of NE. Here, we used Peltier element-controlled incubators and challenged murine hypothalamic mHypoA -2/10 cells with temperature changes of ±1°C. We observed enhanced NE-induced activation of a cAMP-dependent luciferase reporter at 36 compared with 38°C. mRNA analysis and subtype specific antagonists revealed that NE activates ß 2- and ß 3-AR in mHypoA-2/10 cells. Agonist binding to the ß 2-AR was temperature insensitive, but measurements of cytosolic cAMP accumulation revealed an increase in efficacy of 45% ± 27% for NE and of 62% ± 33% for the ß 2-AR-selective agonist salmeterol at 36°C. When monitoring NE-promoted cAMP efflux, we observed an increase in the absolute efflux at 36°C. However, the ratio of exported to cytosolic accumulated cAMP is higher at 38°C. We also stimulated cells with NE at 37°C and measured cAMP degradation at 36 and 38°C afterward. We observed increased cAMP degradation at 38°C, indicating enhanced phosphodiesterase activity at higher temperatures. In line with these data, NE-induced activation of the thyreoliberin promoter was found to be enhanced at 36°C. Overall, we show that physiologic temperature changes fine-tune NE-induced cAMP signaling in hypothalamic cells via ß 2-AR by modulating cAMP degradation and the ratio of intra- and extracellular cAMP. SIGNIFICANCE STATEMENT: Increasing cytosolic cAMP levels by activation of G protein-coupled receptors (GPCR) such as the ß 2-adrenergic receptor (AR) is essential for many body functions. Changes in core body temperature are fundamental and universal factors of mammalian life. This study provides the first data linking physiologically relevant temperature fluctuations to ß 2-AR-induced cAMP signaling, highlighting a so far unappreciated role of body temperature as a modulator of the prototypic class A GPCR.

ß3-adrenoceptor activation exhibits a dual effect on behaviors and glutamate receptor function in the prefrontal cortex.

ß-adrenoceptor (ß-AR), especially the ß1- and ß2-AR subtypes, is known to participate in stress-related behavioral changes. Recently, SR58611A, a brain-penetrant ß3-AR agonist, exhibits anxiolytic- and antidepressant-like effects. In this study, we sought to study the role of SR58611A in behavioral changes and its potential cellular and molecular mechanism in the prefrontal cortex (PFC). We found that rats with SR58611A (1 mg/kg) enhanced PFC-mediated recognition memory, whereas administration of higher dosage of SR58611A (20 mg/kg) caused hyperlocomotion, and exhibited an impairment effect on recognition memory. Electrophysiological data also indicated that SR58611A (1 mg/kg) selectively enhanced NMDA receptor-mediated excitatory postsynaptic currents (EPSC) through interacting with norepinephrine (NE) system and activating ß3-AR, whereas higher dosage of SR58611A (20 mg/kg) reduced both AMPA receptor- and NMDA receptor-mediated EPSC. SR58611A-induced different effects on EPSC linked with the change of the surface expression quantity of NMDA receptor and/or AMPA receptor subunits. Synaptosomal-associated protein 25 (SNAP-25), which is a key soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein involved in incorporation of NMDA receptor to postsynaptic membrane, contributed to SR58611A (1 mg/kg)-induced enhancement of recognition memory and NMDA receptor function. Moreover, SR58611A (1 mg/kg) could rescue repeated stress-induced defect of both recognition memory and NMDA receptor function through a SNAP-25-dependent mechanism. These results provide a potential mechanism underlying the cognitive-enhancing effects of SR58611A (1 mg/kg).

Noradrenaline protects neurons against H2 O2 -induced death by increasing the supply of glutathione from astrocytes via ß3 -adrenoceptor stimulation.

Oxidative stress has been implicated in a variety of neurodegenerative disorders, such as Alzheimer's and Parkinson's disease. Astrocytes play a significant role in maintaining survival of neurons by supplying antioxidants such as glutathione (GSH) to neurons. Recently, we found that noradrenaline increased the intracellular GSH concentration in astrocytes via ß3 -adrenoceptor stimulation. These observations suggest that noradrenaline protects neurons from oxidative stress-induced death by increasing the supply of GSH from astrocytes to neurons via the stimulation of ß3 -adrenoceptor in astrocytes. In the present study, we examined the protective effect of noradrenaline against H2 O2 -induced neurotoxicity using two different mixed cultures: the mixed culture of human astrocytoma U-251 MG cells and human neuroblastoma SH-SY5Y cells, and the mouse primary cerebrum mixed culture of neurons and astrocytes. H2 O2 -induced neuronal cell death was significantly attenuated by pretreatment with noradrenaline in both mixed cultures but not in single culture of SH-SY5Y cells or in mouse cerebrum neuron-rich culture. The neuroprotective effect of noradrenaline was inhibited by SR59230A, a selective ß3 -adrenoceptor antagonist, and CL316243, a selective ß3 -adrenoceptor agonist, mimicked the neuroprotective effect of noradrenaline. DL-buthionine-[S,R]-sulfoximine, a GSH synthesis inhibitor, negated the neuroprotective effect of noradrenaline in both mixed cultures. MK571, which inhibits the export of GSH from astrocytes mediated by multidrug resistance-associated protein 1, also prevented the neuroprotective effect of noradrenaline. These results suggest that noradrenaline protects neurons against H2 O2 -induced death by increasing the supply of GSH from astrocytes via ß3 -adrenoceptor stimulation.

Role of ß3-adrenergic receptor in the modulation of synaptic transmission and plasticity in mouse cerebellar cortex.

Convergent lines of evidence have recently highlighted ß3-adrenoreceptors (ARs) as a potentially critical target in the regulation of nervous and behavioral functions, including memory consolidation, anxiety, and depression. Nevertheless, the role of ß3-ARs in the cerebellum has been never investigated. To address this issue, we first examined the effects of pharmacological manipulation of ß3-ARs on motor learning in mice. We found that blockade of ß3-ARs by SR 59230A impaired the acquisition of the rotarod task with no effect on general locomotion. Since the parallel fiber-Purkinje cell (PF-PC) synapse is considered to be the main cerebellar locus of motor learning, we assessed ß3-AR modulatory action on this synapse as well as its expression in cerebellar slices. We demonstrate, for the first time, a strong expression of ß3-ARs on Purkinje cell soma and dendrites. In addition, whole-cell patch-clamp recordings revealed that bath application of ß3-AR agonist CL316,243 depressed the PF-PC excitatory postsynaptic currents via a postsynaptic mechanism mediated by the PI3K signaling pathway. Application of CL316,243 also interfered with the expression of PF long-term potentiation, whereas SR 59230A prevented the induction of LTD at PF-PC synapse. These results underline the critical role of ß3-AR on cerebellar synaptic transmission and plasticity and provide a new mechanism for adrenergic modulation of motor learning.

ß3-Adrenoceptors as Putative Regulator of Immune Tolerance in Cancer and Pregnancy.

Understanding the mechanisms of immune tolerance is currently one of the most important challenges of scientific research. Pregnancy affects the immune system balance, leading the host to tolerate embryo alloantigens. Previous reports demonstrated that ß-adrenergic receptor (ß-AR) signaling promotes immune tolerance by modulation of NK and Treg, mainly through the activation of ß2-ARs, but recently we have demonstrated that also ß3-ARs induce an immune-tolerant phenotype in mice bearing melanoma. In this report, we demonstrate that ß3-ARs support host immune tolerance in the maternal microenvironment by modulating the same immune cells populations as recently demonstrated in cancer. Considering that ß3-ARs are modulated by oxygen levels, we hypothesize that hypoxia, through the upregulation of ß3-AR, promotes the biological shift toward a tolerant immunophenotype and that this is the same trick that embryo and cancer use to create an aura of immune-tolerance in a competent immune environment. This study confirms the analogies between fetal development and tumor progression and suggests that the expression of ß3-ARs represents one of the strategies to induce fetal and tumor immune tolerance.

Perivascular adipose tissue phenotype and sepsis vascular dysfunction: Differential contribution of NO, ROS and beta 3-adrenergic receptor.

AIMS: Vascular dysfunction plays a key role in sepsis but the role of perivascular adipose tissue (PVAT) in this condition is relatively unknown. MAIN METHODS: Sepsis was induced by cecal ligation and puncture (CLP). The responses of the aorta and superior mesenteric artery to norepinephrine in the presence or absence of PVAT were evaluated. Fluorescent probes measured the production of nitric oxide (NO) and reactive oxygen species (ROS). NO synthases (NOS) and ß3-adrenoceptor expression were detected by immunofluorescence and S-nitrosylation by the biotin switch assay. KEY FINDINGS: Aorta and superior mesenteric arteries from septic animals with intact PVAT showed a worsened response to the vasoconstrictor compared to vessels without PVAT. PVAT from the aorta (APVAT) produced NO and ROS whereas PVAT from the superior mesenteric artery (MPVAT) produced only ROS. Septic APVAT exhibited a higher density of NOS-1 and NOS-3. S-nitrosylation was found in APVAT. Donor (PVAT obtained from normal or septic rats):Host (normal vessel without PVAT) experiments showed that L-NAME, ODQ and ß3-adrenergic receptor antagonist blocked the septic APVAT anti-contractile effect. None of these compounds affected MPVAT; tempol, but not apocynin, blocked its anti-contractile effect. SIGNIFICANCE: PVAT contributes to the anti-contractile effect in the aorta and mesenteric artery of septic rats through different pathways. ß3-Adrenergic receptor and NO appear to be key mediators of this effect in APVAT, but not in MPVAT where ROS seem to be a relevant mediator. Therefore, PVAT is a relevant player of sepsis vascular dysfunction.

Combating Obesity With Thermogenic Fat: Current Challenges and Advancements.

Brown fat and beige fat are known as thermogenic fat due to their contribution to non-shivering thermogenesis in mammals following cold stimulation. Beige fat is unique due to its origin and its development in white fat. Subsequently, both brown fat and beige fat have become viable targets to combat obesity. Over the last few decades, most therapeutic strategies have been focused on the canonical pathway of thermogenic fat activation via the ß3-adrenergic receptor (AR). Notwithstanding, administering ß3-AR agonists often leads to side effects including hypertension and particularly cardiovascular disease. It is thus imperative to search for alternative therapeutic approaches to combat obesity. In this review, we discuss the current challenges in the field with respect to stimulating brown/beige fat thermogenesis. Additionally, we include a summary of other newly discovered pathways, including non-AR signaling- and non-UCP1-dependent mechanisms, which could be potential targets for the treatment of obesity and its related metabolic diseases.

Real-time monitoring of cAMP in brown adipocytes reveals differential compartmentation of ß1 and ß3-adrenoceptor signalling.

OBJECTIVE: 3',5'-Cyclic adenosine monophosphate (cAMP) is a central second messenger governing brown adipocyte differentiation and function. ß-adrenergic receptors (ß-ARs) stimulate adenylate cyclases which produce cAMP. Moreover, cyclic nucleotide levels are tightly controlled by phosphodiesterases (PDEs), which can generate subcellular microdomains of cAMP. Since the spatio-temporal organisation of the cAMP signalling pathway in adipocytes is still unclear, we sought to monitor real-time cAMP dynamics by live cell imaging in pre-mature and mature brown adipocytes. METHODS: We measured the real-time dynamics of cAMP in murine pre-mature and mature brown adipocytes during stimulation of individual ß-AR subtypes, as well as its regulation by PDEs using a Förster Resonance Energy Transfer based biosensor and pharmacological tools. We also correlated these data with ß-AR stimulated lipolysis and analysed the expression of ß-ARs and PDEs in brown adipocytes using qPCR and immunoblotting. Furthermore, subcellular distribution of PDEs was studied using cell fractionation and immunoblots. RESULTS: Using pre-mature and mature brown adipocytes isolated from transgenic mice expressing a highly sensitive cytosolic biosensor Epac1-camps, we established real-time measurements of cAMP responses. PDE4 turned out to be the major PDE regulating cytosolic cAMP in brown preadipocytes. Upon maturation, PDE3 gets upregulated and contributes with PDE4 to control ß1-AR-induced cAMP. Unexpectedly, ß3-AR initiated cAMP is resistant to increased PDE3 protein levels and simultaneously, the control of this microdomain by PDE4 is reduced upon brown adipocyte maturation. Therefore we postulate the existence of distinct cAMP pools in brown adipocytes. One cAMP pool is formed by ß1-AR associated with PDE3 and PDE4, while another pool is centred around ß3-AR and is much less controlled by these PDEs. Functionally, lower control of ß3-AR initiated cAMP by PDE3 and PDE4 facilitates brown adipocyte lipolysis, while lipolysis activated by ß1-AR and is under tight control of PDE3 and PDE4. CONCLUSIONS: We have established a real-time live cell imaging approach to analyse brown adipocyte cAMP dynamics in real-time using a cAMP biosensor. We showed that during the differentiation from pre-mature to mature murine brown adipocytes, there was a change in PDE-dependent compartmentation of ß1-and ß3-AR-initiated cAMP responses by PDE3 and PDE4 regulating lipolysis.

Noradrenaline-Induced Relaxation of Urinary Bladder Smooth Muscle Is Primarily Triggered through the ß3-Adrenoceptor in Rats.

ß-Adrenoceptors are subclassified into 3 subtypes (ß1-ß3). Among these, ß3-adrenoceptors are present in various types of smooth muscle and are believed to play a role in relaxation responses of these muscles. ß3-Adrenoceptors are also present in urinary bladder smooth muscle (UBSM), although their expression varies depending on the animal species. To date, there has been little information available about the endogenous ligand that stimulates ß3-adrenoceptors to produce relaxation responses in UBSM. In this study, to determine whether noradrenaline is a ligand of UBSM ß3-adrenoceptors, noradrenaline-induced relaxation was analyzed pharmacologically using rat UBSM. We also assessed whether noradrenaline metabolites were ligands in UBSM. In isolated rat urinary bladder tissues, mRNAs for ß1-, ß2-, and ß3-adrenoceptors were detected using RT-PCR. In UBSM preparations contracted with methacholine (3 × 10-5 M), noradrenaline-induced relaxation was not inhibited by the following antagonists: atenolol (10-6 M; selective ß1-adrenoceptor antagonist), ICI-118,551 (3 × 10-8 M; selective ß2-adrenoceptor antagonist), propranolol (10-7 M; non-selective ß-adrenoceptor antagonist), and bupranolol (10-7 M; non-selective ß-adrenoceptor antagonist). In the presence of propranolol (10-6 M), noradrenaline-induced relaxation was competitively inhibited by bupranolol (3 × 10-7-3 × 10-6 M) or SR59230A (10-7-10-6 M; selective ß3-adrenoceptor antagonist), with their pA2 values calculated to be 6.64 and 7.27, respectively. None of the six noradrenaline metabolites produced significant relaxation of methacholine-contracted UBSM. These findings suggest that noradrenaline, but not its metabolites, is a ligand for ß3-adrenoceptors to produce relaxation responses of UBSM in rats.

Everything You Always Wanted to Know about ß3-AR * (* But Were Afraid to Ask).

The beta-3 adrenergic receptor (ß3-AR) is by far the least studied isotype of the beta-adrenergic sub-family. Despite its study being long hampered by the lack of suitable animal and cellular models and inter-species differences, a substantial body of literature on the subject has built up in the last three decades and the physiology of ß3-AR is unraveling quickly. As will become evident in this work, ß3-AR is emerging as an appealing target for novel pharmacological approaches in several clinical areas involving metabolic, cardiovascular, urinary, and ocular disease. In this review, we will discuss the most recent advances regarding ß3-AR signaling and function and summarize how these findings translate, or may do so, into current clinical practice highlighting ß3-AR's great potential as a novel therapeutic target in a wide range of human conditions.

Depletion of ß3-adrenergic receptor induces left ventricular diastolic dysfunction via potential regulation of energy metabolism and cardiac contraction.

Left ventricular diastolic dysfunction (LVDD) is a central perturbation in heart failure with preserved ejection fraction, and there are currently no effective remedies to improve LVDD in clinical practice. The ß3-adrenergic receptor (ADRB3) was reported to play protective effects on inhibiting myocardial fibrosis in response to hemodynamic stress. However, the effects of ADRB3 on LVDD and its underlying mechanisms are still undefined. In the current study, the role of ADRB3 in LVDD was identified in ADRB3-knockout mice. Echocardiography parameters showed that depletion of ADRB3 had little effect on cardiac systolic function but obviously led to cardiac diastolic dysfunction in vivo. Proteomics (including the global proteome, phosphorylated and acetylated proteome) and bioinformatics analysis (including GO analysis, KEGG pathway analysis, GO-Tree network, Pathway-Act network, and protein-protein interaction network) were performed on cardiac specimens of ADRB3-KO mice and wild-type mice. The results showed that the cardiac energy metabolism (especially the citrate cycle), actin cytoskeleton organization, and cardiac muscle contraction (related to mitogen-activated protein kinase, toll-like receptor, and ErbB signalling pathway) were potential core mechanisms underlying ADRB3-KO-induced LVDD. In addition, the protein-protein interaction network indicated that the core proteins associated with ADRB3-KO-induced LVDD were FGG, ALDH1A1, FGA, APOC3, SLC4A1, SERPINF2, HP, CTNNB1, and TKT. In conclusion, the absence of ADRB3 leads to LVDD, which is potentially associated with the regulation of cardiac energy metabolism, actin cytoskeleton organization, and cardiac muscle contraction.

Physiology and pathophysiology of the ß3-adrenergic receptor.

The ß3-adrenergic receptor (ß3-AR) is an important regulator of various physiological functions, such as thermogenesis in brown adipose tissue, lipolysis in white adipose tissue, negative inotropic effect in cardiomyocyte, and relaxation in blood vessel. The activation of ß3-AR by its agonists is shown to have metabolic (antiobesity and antidiabetic) and cardiovascular effects in animal models, highlighting ß3-AR as a potential therapeutic target in the treatment of several human diseases. Moreover, a substantial number of studies performed on different populations have identified some ß3-AR polymorphic variants associated with obesity, diabetes, cardiovascular diseases, and other disorders. The clinical phenotypes and functional characteristics of these variants provide insights into potential pathophysiological roles of ß3-AR in the development of these diseases.

Dual cholinergic signals regulate daily migration of hematopoietic stem cells and leukocytes.

Hematopoietic stem and progenitor cells (HSPCs) and leukocytes circulate between the bone marrow (BM) and peripheral blood following circadian oscillations. Autonomic sympathetic noradrenergic signals have been shown to regulate HSPC and leukocyte trafficking, but the role of the cholinergic branch has remained unexplored. We have investigated the role of the cholinergic nervous system in the regulation of day/night traffic of HSPCs and leukocytes in mice. We show here that the autonomic cholinergic nervous system (including parasympathetic and sympathetic) dually regulates daily migration of HSPCs and leukocytes. At night, central parasympathetic cholinergic signals dampen sympathetic noradrenergic tone and decrease BM egress of HSPCs and leukocytes. However, during the daytime, derepressed sympathetic noradrenergic activity causes predominant BM egress of HSPCs and leukocytes via ß3-adrenergic receptor. This egress is locally supported by light-triggered sympathetic cholinergic activity, which inhibits BM vascular cell adhesion and homing. In summary, central (parasympathetic) and local (sympathetic) cholinergic signals regulate day/night oscillations of circulating HSPCs and leukocytes. This study shows how both branches of the autonomic nervous system cooperate to orchestrate daily traffic of HSPCs and leukocytes.

α1- and ß3-Adrenergic Receptor-Mediated Mesolimbic Homeostatic Plasticity Confers Resilience to Social Stress in Susceptible Mice.

BACKGROUND: Homeostatic plasticity in mesolimbic dopamine (DA) neurons plays an essential role in mediating resilience to social stress. Recent evidence implicates an association between stress resilience and projections from the locus coeruleus (LC) to the ventral tegmental area (VTA) (LC→VTA) DA system. However, the precise circuitry and molecular mechanisms of the homeostatic plasticity in mesolimbic DA neurons mediated by the LC→VTA circuitry, and its role in conferring resilience to social defeat stress, have not been described. METHODS: In a well-established chronic social defeat stress model of depression, using projection-specific electrophysiological recordings and optogenetic, pharmacological, and molecular profiling techniques, we investigated the functional role and molecular basis of an LC→VTA circuit in conferring resilience to social defeat stress. RESULTS: We found that LC neurons projecting to the VTA exhibit enhanced firing activity in resilient, but not susceptible, mice. Optogenetically mimicking this firing adaptation in susceptible mice reverses their depression-related behaviors, and induces reversal of cellular hyperactivity and homeostatic plasticity in VTA DA neurons projecting to the nucleus accumbens. Circuit-specific molecular profiling studies reveal that α1- and ß3-adrenergic receptors are highly expressed in VTA→nucleus accumbens DA neurons. Pharmacologically activating these receptors induces similar proresilient effects at the ion channel and cellular and behavioral levels, whereas antagonizing these receptors blocks the proresilient effect of optogenetic activation of LC→VTA circuit neurons in susceptible mice. CONCLUSIONS: These findings reveal a key role of the LC→VTA circuit in mediating homeostatic plasticity in stress resilience and reveal α1- and ß3-adrenergic receptors as new molecular targets for therapeutically promoting resilience.

Adrenergic receptor ß3 is involved in the memory consolidation process in mice.

Attention and emotion have a positive impact on memory formation, which is related to the activation of the noradrenergic system in the brain. The hippocampus and amygdala are fundamental structures in memory acquisition, which is modulated by noradrenaline through the noradrenergic receptors. Pharmacological studies suggest that memory acquisition depends on the action of both the ß3 (ß3-AR) and ß2 (ß2-AR) receptor subtypes. However, the use of animal models with specific knockout for the ß3-AR receptor only (ß3-ARKO) allows researchers to more accurately assess its role in memory formation processes. In the present study, we evaluated short- and long-term memory acquisition capacity in ß3-ARKO mice and wild-type mice at approximately 60 days of age. The animals were submitted to the open field test, the elevated plus maze, object recognition, and social preference. The results showed that the absence of the ß3-AR receptor caused no impairment in locomotion and did not cause anxious behavior, but it caused significant impairment of short- and long-term memory compared to wild-type animals. We also evaluated the expression of genes involved in memory consolidation. The mRNA levels for GLUT3, a glucose transporter expressed in the central nervous system, were significantly reduced in the amygdala, but not in the hippocampus of the ß3-ARKO animals. Our results showed that ß3-AR was involved in the process of acquisition of declarative memory, and its action may be due to the facilitation of glucose absorption in the amygdala.

Sustained stimulation of ß2- and ß3-adrenergic receptors leads to persistent functional pain and neuroinflammation.

Functional pain syndromes, such as fibromyalgia and temporomandibular disorder, are associated with enhanced catecholamine tone and decreased levels of catechol-O-methyltransferase (COMT; an enzyme that metabolizes catecholamines). Consistent with clinical syndromes, our lab has shown that sustained 14-day delivery of the COMT inhibitor OR486 in rodents results in pain at multiple body sites and pain-related volitional behaviors. The onset of COMT-dependent functional pain is mediated by peripheral ß2- and ß3-adrenergic receptors (ß2- and ß3ARs) through the release of the pro-inflammatory cytokines tumor necrosis factor α (TNFα), interleukin-1ß (IL-1ß), and interleukin-6 (IL-6). Here, we first sought to investigate the role of ß2- and ß3ARs and downstream mediators in the maintenance of persistent functional pain. We then aimed to characterize the resulting persistent inflammation in neural tissues (neuroinflammation), characterized by activated glial cells and phosphorylation of the mitogen-activated protein kinases (MAPKs) p38 and extracellular signal-regulated kinase (ERK). Separate groups of rats were implanted with subcutaneous osmotic mini-pumps to deliver OR486 (15 mg/kg/day) or vehicle for 14 days. The ß2AR antagonist ICI118551 and ß3AR antagonist SR59230A were co-administrated subcutaneously with OR486 or vehicle either on day 0 or day 7. The TNFα inhibitor Etanercept, the p38 inhibitor SB203580, or the ERK inhibitor U0126 were delivered intrathecally following OR486 cessation on day 14. Behavioral responses, pro-inflammatory cytokine levels, glial cell activation, and MAPK phosphorylation were measured over the course of 35 days. Our results demonstrate that systemic delivery of OR486 leads to mechanical hypersensitivity that persists for at least 3 weeks after OR486 cessation. Corresponding increases in spinal TNFα, IL-1ß, and IL-6 levels, microglia and astrocyte activation, and neuronal p38 and ERK phosphorylation were observed on days 14-35. Persistent functional pain was alleviated by systemic delivery of ICI118551 and SR59230A beginning on day 0, but not day 7, and by spinal delivery of Etanercept or SB203580 beginning on day 14. These results suggest that peripheral ß2- and ß3ARs drive persistent COMT-dependent functional pain via increased activation of immune cells and production of pro-inflammatory cytokines, which promote neuroinflammation and nociceptor activation. Thus, therapies that resolve neuroinflammation may prove useful in the management of functional pain syndromes.

Adrenergic receptor ß3 is involved in the memory consolidation process in mice

Attention and emotion have a positive impact on memory formation, which is related to the activation of the noradrenergic system in the brain. The hippocampus and amygdala are fundamental structures in memory acquisition, which is modulated by noradrenaline through the noradrenergic receptors. Pharmacological studies suggest that memory acquisition depends on the action of both the β3 (β3-AR) and β2 (β2-AR) receptor subtypes. However, the use of animal models with specific knockout for the β3-AR receptor only (β3-ARKO) allows researchers to more accurately assess its role in memory formation processes. In the present study, we evaluated short- and long-term memory acquisition capacity in β3-ARKO mice and wild-type mice at approximately 60 days of age. The animals were submitted to the open field test, the elevated plus maze, object recognition, and social preference. The results showed that the absence of the β3-AR receptor caused no impairment in locomotion and did not cause anxious behavior, but it caused significant impairment of short- and long-term memory compared to wild-type animals. We also evaluated the expression of genes involved in memory consolidation. The mRNA levels for GLUT3, a glucose transporter expressed in the central nervous system, were significantly reduced in the amygdala, but not in the hippocampus of the β3-ARKO animals. Our results showed that β3-AR was involved in the process of acquisition of declarative memory, and its action may be due to the facilitation of glucose absorption in the amygdala.

ß3 adrenergic receptor activation relaxes human corpus cavernosum and penile artery through a hydrogen sulfide/cGMP-dependent mechanism.

Erectile function is a widely accepted indicator of systemic endothelial activity since from a clinical standpoint erectile dysfunction (ED) often precedes cardiovascular events. Recently it has been described a potential role for ß3 adrenoceptor in cardiovascular diseases emphasizing a possible development of new drugs. ß3 adrenoceptor stimulation relaxes human corpus cavernosum (HCC) strips in cyclic guanosine monophosphate (cGMP)-dependent and endothelium/nitric oxide (NO)-independent manner. Hydrogen sulfide (H2S), along with NO, is another gaseous molecule involved in cardiovascular system and as a consequence also in penile erection. Cystathionine-ß-synthase (CBS) and cystathionine-γ-lyase (CSE), the enzymes mainly responsible for H2S biosynthesis, are constitutively expressed in HCC. CSE rather than CBS is more abundant in human penile tissue. Herein we investigated the involvement of H2S pathway in ß3 adrenoceptor-induced relaxation in HCC and penile artery. Penile artery expresses both CSE and ß3 adrenoceptor. BRL37344, a ß3 selective agonist, relaxed HCC strips and penile artery rings and this effect was significantly reduced by CSE inhibition. Incubation of HCC and penile artery homogenate with BRL37344 significantly increased H2S production. This effect was significantly reduced by the inhibition of either CSE or ß3 adrenoceptor. Finally, the BRL37344-induced increase in cGMP was reduced by CSE inhibition in both tissues. Thus, BRL37344-induced relaxation in HCC and penile artery occurs in a H2S/cGMP-dependent manner. In conclusion, ß3/H2S/cGMP pathway can act as an alternative to NO. Since about 15% of patients do not respond to phosphodiesterase-5 inhibitors, ß3 agonists could represent a therapeutic alternative or a useful adjuvant therapy to treat these patients.

[Effects 'of ß3 adrenoceptors on the contractility of rat thoracic aorta smooth muscle and the mechanism].

OBJECTIVE: To observe the effect of ß3adrenoceptors (ß3-AR) activation on rat thoracic aorta smooth muscle contractility and the possible related mechanism. METHODS: The endothelium removed thoracic aorta was pre-contracted with 30 mmol/L KCl physiological saline solution (PSS). Then the tension of the thoracic aorta was recorded in presence of BRL37344 (BRL) to determine the action of ß3-AR. The tension of the thoracic aorta was also recorded in the presence of Propranolol (PRA), SR59230A (SR), L-NNA, H-89 and Iberiotoxin (IBTX) respectively to reveal the underling mechanism of ß3-AR activation on rat vascular smooth muscle. Immunohistochemistry was adopted to confirm the existence and the distribution of ß3-AR in rat thoracic aorta. RESULTS: The results showed that: (1) The thoracic aorta was relaxed by ß3-AR activation, with a relaxation percentage of (10.59 ± 0.79). (2) ß3-AR was expressed in both endothelial and smooth muscle layer in thoracic aorta sections of rats. (3) PRA did not block the effect of BRL on the thoracic aorta. The relaxation actions of BRL could be antagonized by pre-incubating the thoracic aorta with SR. (4) L-NNA (a NOS inhibitor) and H-89 (a PKA inhibitor) reversed the relaxation effect of BRL on vascular smooth muscle. (5) The effect of BRL was decreased after application of Ibriotoxin (IBTX), a large conductance calcium dependent potassium channel blocker. CONCLUSION: The results confirmed that activation of ß3-AR led to relaxation of thoracic aorta smooth muscle. The relaxation action of ß3-AR on smooth muscle of rat thoracic aorta was related to activation of NOS and PKA signaling pathway. Large conductance Ca²âº-K⁺ channels were involved in the relaxation action of ß3-AR activation on rat thoracic aorta smooth muscle.