Cerebrovascular and cardiovascular autonomic regulation in sickle cell patients with white matter lesions.

BACKGROUND AND PURPOSE: White matter lesions (WMLs) are frequent in sickle cell disease (SCD), with a prevalence described to be as high as 53% by age 30. Cerebrovascular regulation and cardiovascular autonomic regulation, more specifically the sympatho-vagal balance, can be altered in SCD. In this study the association between WMLs, cerebrovascular regulation and sympatho-vagal balance was assessed in SCD patients. METHODS AND RESULTS: Sickle cell disease patients with no history of stroke were prospectively evaluated for cerebrovascular reactivity using the breath-holding test (BHT), the sympatho-vagal balance (ratio low frequency/high frequency [HF]) using heart rate variability parameters and cerebral autoregulation in the time domain using correlation index Mx, and arterial cerebral compliance based on continuous assessment of cerebral blood flow velocities using transcranial Doppler ultrasound and arterial blood pressure with photo-plethysmography. WMLs were assessed with magnetic resonance imaging using Fazekas score grading and the presence of lacunes. Forty-one patients (F/M 25/16) were included. Median age was 37.5 years (19-65). Twenty-nine (70.7%) patients had SS genotype. Eleven patients had WMLs (26.8%). Patients with WMLs were significantly older (p < 0.001), had a lower HF (p < 0.005) and an impaired cerebral arterial compliance (p < 0.014). The receiver operating curve for the regression model including age and HF showed a higher area under the curve compared to age alone (0.946 vs. 0.876). BHT and Mx did not significantly differ between the two groups. CONCLUSIONS: Lower parasympathetic activity and impaired cerebral arterial compliance were associated with WMLs in adults with SCD. This could potentially yield to a better understanding of pathophysiological parameters leading to premature cerebrovascular ageing in SCD.

Platelet P2Y1 receptor exhibits constitutive G protein signaling and ß-arrestin 2 recruitment.

BACKGROUND: Purinergic P2Y1 and P2Y12 receptors (P2Y1-R and P2Y12-R) are G protein-coupled receptors (GPCR) activated by adenosine diphosphate (ADP) to mediate platelet activation, thereby playing a pivotal role in hemostasis and thrombosis. While P2Y12-R is the major target of antiplatelet drugs, no P2Y1-R antagonist has yet been developed for clinical use. However, accumulating data suggest that P2Y1-R inhibition would ensure efficient platelet inhibition with minimal effects on bleeding. In this context, an accurate characterization of P2Y1-R antagonists constitutes an important preliminary step. RESULTS: Here, we investigated the pharmacology of P2Y1-R signaling through Gq and ß-arrestin pathways in HEK293T cells and in mouse and human platelets using highly sensitive resonance energy transfer-based technologies (BRET/HTRF). We demonstrated that at basal state, in the absence of agonist ligand, P2Y1-R activates Gq protein signaling in HEK293T cells and in mouse and human platelets, indicating that P2Y1-R is constitutively active in physiological conditions. We showed that P2Y1-R also promotes constitutive recruitment of ß-arrestin 2 in HEK293T cells. Moreover, the P2Y1-R antagonists MRS2179, MRS2279 and MRS2500 abolished the receptor dependent-constitutive activation, thus behaving as inverse agonists. CONCLUSIONS: This study sheds new light on P2Y1-R pharmacology, highlighting for the first time the existence of a constitutively active P2Y1-R population in human platelets. Given the recent interest of P2Y12-R constitutive activity in patients with diabetes, this study suggests that modification of constitutive P2Y1-R signaling might be involved in pathological conditions, including bleeding syndrome or high susceptibility to thrombotic risk. Thus, targeting platelet P2Y1-R constitutive activation might be a promising and powerful strategy for future antiplatelet therapy.

Impact of the COVID-19 pandemic on clinical autonomic practice in Europe A survey of the European Academy of Neurology (EAN) and the European Federation of Autonomic Societies (EFAS).

OBJECTIVE: To investigate the impact of the coronavirus-disease-2019 (COVID-19) pandemic on European clinical autonomic practice. METHODS: Eighty-four neurology-driven or interdisciplinary autonomic centers in 22 European countries were invited to fill in a web-based survey between September and November 2021. RESULTS: Forty-six centers completed the survey (55%). During the first pandemic year, the number of performed tilt-table tests, autonomic outpatient and inpatient visits decreased respectively by 50%, 45% and 53%, and every-third center reported major adverse events due to postponed examinations or visits. The most frequent newly-diagnosed or worsened cardiovascular autonomic disorders after COVID-19 infection included postural orthostatic tachycardia syndrome (POTS), orthostatic hypotension, and recurrent vasovagal syncope, deemed likely related to the infection by ≥50% of the responders. Forty-seven percent of the responders also reported about people with new-onset of orthostatic intolerance, but negative tilt-table findings, and 16% about people with psychogenic pseudosyncope after COVID-19. Most patients were treated non-pharmacologically and symptomatic recovery at follow-up was observed in ≥45% of cases. By contrast, low frequencies of newly-diagnosed cardiovascular autonomic disorders following COVID-19 vaccination were reported, most frequently POTS and recurrent vasovagal syncope, and most of the responders judged a causal association unlikely. Non-pharmacological measures were the preferred treatment choice, with 50-100% recovery rates at follow-up. CONCLUSIONS: Cardiovascular autonomic disorders may develop or worsen following a COVID-19 infection, while the association with COVID-19 vaccines remains controversial. Despite the severe pandemic impact on European clinical autonomic practice, a specialized diagnostic work-up was pivotal to identify non-autonomic disorders in people with post-COVID-19 orthostatic complaints.

EFAS/EAN survey on the influence of the COVID-19 pandemic on European clinical autonomic education and research.

PURPOSE: To understand the influence of the coronavirus disease 2019 (COVID-19) pandemic on clinical autonomic education and research in Europe. METHODS: We invited 84 European autonomic centers to complete an online survey, recorded the pre-pandemic-to-pandemic percentage of junior participants in the annual congresses of the European Federation of Autonomic Societies (EFAS) and European Academy of Neurology (EAN) and the pre-pandemic-to-pandemic number of PubMed publications on neurological disorders. RESULTS: Forty-six centers answered the survey (55%). Twenty-nine centers were involved in clinical autonomic education and experienced pandemic-related didactic interruptions for 9 (5; 9) months. Ninety percent (n = 26/29) of autonomic educational centers reported a negative impact of the COVID-19 pandemic on education quality, and 93% (n = 27/29) established e-learning models. Both the 2020 joint EAN-EFAS virtual congress and the 2021 (virtual) and 2022 (hybrid) EFAS and EAN congresses marked higher percentages of junior participants than in 2019. Forty-one respondents (89%) were autonomic researchers, and 29 of them reported pandemic-related trial interruptions for 5 (2; 9) months. Since the pandemic begin, almost half of the respondents had less time for scientific writing. Likewise, the number of PubMed publications on autonomic topics showed the smallest increase compared with other neurological fields in 2020-2021 and the highest drop in 2022. Autonomic research centers that amended their trial protocols for telemedicine (38%, n = 16/41) maintained higher clinical caseloads during the first pandemic year. CONCLUSIONS: The COVID-19 pandemic had a substantial negative impact on European clinical autonomic education and research. At the same time, it promoted digitalization, favoring more equitable access to autonomic education and improved trial design.

Ambulatory blood pressure and drug treatment for orthostatic hypotension as predictors of mortality in patients with multiple system atrophy.

OBJECTIVES: Multiple system atrophy (MSA) is a rare fatal neurodegenerative disease characterized by parkinsonism, cerebellar ataxia and autonomic failure. This study was aimed at investigating possible associations between mortality, 24-h blood pressure (BP) level and variability, and drug treatments for orthostatic hypotension (OH) in MSA patients. METHODS: A total of 129 patients followed at the French Reference Center for MSA who underwent routine 24-h ambulatory BP monitoring were included. Unified MSA Rating Scale (UMSARS) scores, drug treatments and the occurrence and cause of death were recorded. RESULTS: Seventy patients died during follow-up (2.9 ± 1.8 years), mainly from terminal illness, pulmonary or sudden death. Multivariate Cox regression analysis, after adjustment for gender, disease duration and severity (UMSARS I+II score), showed that increased daytime systolic BP variability, OH severity and OH drug treatment were independently correlated with mortality. OH treatment was associated with the risk of cardiac causes and/or sudden death (p = 0.01). In a fully adjusted model, male gender [(female vs. male) hazard ratio (HR) 0.56, 95% CI 0.34-0.94, p = 0.03], UMSARS I+II score (HR 1.04, 95% CI 1.02-1.06, p < 0.01), systolic BP daytime variability (HR 3.66, 95% CI 1.46-9.17, p < 0.01) and OH treatment (HR: 2.13, 95% CI 1.15-3.94, p = 0.02) predicted mortality. CONCLUSIONS: Increased daytime BP variability and OH treatment were predictive of mortality in patients with MSA, independently from disease severity. Further studies are required to assess if these associations are explained by more severe autonomic dysfunction or if OH treatment exposes per se to a specific risk in this population.

Clinical autonomic nervous system laboratories in Europe: A joint survey of the European Academy of Neurology and the European Federation of Autonomic Societies: A joint survey of the European Academy of Neurology and the European Federation of Autonomic Societies.

BACKGROUND AND PURPOSE: Disorders of the autonomic nervous system (ANS) are common conditions, but it is unclear whether access to ANS healthcare provision is homogeneous across European countries. The aim of this study was to identify neurology-driven or interdisciplinary clinical ANS laboratories in Europe, describe their characteristics and explore regional differences. METHODS: We contacted the European national ANS and neurological societies, as well as members of our professional network, to identify clinical ANS laboratories in each country and invite them to answer a web-based survey. RESULTS: We identified 84 laboratories in 22 countries and 46 (55%) answered the survey. All laboratories perform cardiovascular autonomic function tests, and 83% also perform sweat tests. Testing for catecholamines and autoantibodies are performed in 63% and 56% of laboratories, and epidermal nerve fiber density analysis in 63%. Each laboratory is staffed by a median of two consultants, one resident, one technician and one nurse. The median (interquartile range [IQR]) number of head-up tilt tests/laboratory/year is 105 (49-251). Reflex syncope and neurogenic orthostatic hypotension are the most frequently diagnosed cardiovascular ANS disorders. Thirty-five centers (76%) have an ANS outpatient clinic, with a median (IQR) of 200 (100-360) outpatient visits/year; 42 centers (91%) also offer inpatient care (median 20 [IQR 4-110] inpatient stays/year). Forty-one laboratories (89%) are involved in research activities. We observed a significant difference in the geographical distribution of ANS services among European regions: 11 out of 12 countries from North/West Europe have at least one ANS laboratory versus 11 out of 21 from South/East/Greater Europe (p = 0.021). CONCLUSIONS: This survey highlights disparities in the availability of healthcare services for people with ANS disorders across European countries, stressing the need for improved access to specialized care in South, East and Greater Europe.

Deciphering biased inverse agonism of cangrelor and ticagrelor at P2Y12 receptor.

P2Y12 receptor (P2Y12-R) is one of the major targets for drug inhibiting platelet aggregation in the treatment/prevention of arterial thrombosis. However, the clinical use of P2Y12-R antagonists faces some limitations, such as a delayed onset of action (clopidogrel) or adverse effect profile (ticagrelor, cangrelor), justifying the development of a new generation of P2Y12-R antagonists with a better clinical benefit-risk balance. Although the recent concept of biased agonism offers the possibility to alleviate undesirable adverse effects while preserving therapeutic outcomes, it has never been explored at P2Y12-R. For the first time, using highly sensitive BRET2-based probes, we accurately delineated biased ligand efficacy at P2Y12-R in living HEK293T cells on G protein activation and downstream effectors. We demonstrated that P2Y12-R displayed constitutive Gi/o-dependent signaling that is impaired by the R122C mutation, previously associated with a bleeding disorder. More importantly, we reported the biased inverse agonist efficacy of cangrelor and ticagrelor that could underlie their clinical efficacy. Our study points out that constitutive P2Y12-R signaling is a normal feature of the receptor that might be essential for platelets to respond faster to a vessel injury. From a therapeutic standpoint, our data suggest that the beneficial advantages of antiplatelet drugs might be more related to inverse agonism at P2Y12-R than to antagonism of ADP-mediated signaling. In the future, deciphering P2Y12-R constitutive activity should allow the discovery of more selective biased P2Y12-R blockers demonstrating therapeutic advantages over classical antiplatelet drugs by improving therapeutic outcomes and concomitantly relieving undesirable adverse effects.

Biophysical properties of cardiomyocyte surface explored by multiparametric AFM.

PeakForce Quantitative Nanomechanical Mapping (PeakForce QNM) multiparametric AFM mode was adapted to qualitative and quantitative study of the lateral membrane of cardiomyocytes (CMs), extending this powerful mode to the study of soft cells. On living CM, PeakForce QNM depicted the crests and hollows periodic alternation of cell surface architecture previously described using AFM Force Volume (FV) mode. PeakForce QNM analysis provided better resolution in terms of pixel number compared to FV mode and reduced acquisition time, thus limiting the consequences of spontaneous living adult CM dedifferentiation once isolated from the cardiac tissue. PeakForce QNM mode on fixed CMs clearly visualized subsarcolemmal mitochondria (SSM) and their loss following formamide treatment, concomitant with the interfibrillar mitochondria climbing up and forming heaps at the cell surface. Interestingly, formamide-promoted SSM loss allowed visualization of the sarcomeric apparatus ultrastructure below the plasma membrane. High PeakForce QNM resolution led to better contrasted mechanical maps than FV mode and provided correlation between adhesion, dissipation, mechanical and topographical maps. Modified hydrophobic AFM tip enhanced contrast on adhesion and dissipation maps and suggested that CM surface crests and hollows exhibit distinct chemical properties. Finally, two-dimensional Fast Fourier Transform to objectively quantify AFM maps allowed characterization of periodicity of both sarcomeric Z-line and M-band. Overall, this study validated PeakForce QNM as a valuable and innovative mode for the exploration of living and fixed CMs. In the future, it could be applied to depict cell membrane architectural, mechanical and chemical defects as well as sarcomeric abnormalities associated with cardiac diseases.

Dual agonist occupancy of AT1-R-α2C-AR heterodimers results in atypical Gs-PKA signaling.

Hypersecretion of norepinephrine (NE) and angiotensin II (AngII) is a hallmark of major prevalent cardiovascular diseases that contribute to cardiac pathophysiology and morbidity. Herein, we explore whether heterodimerization of presynaptic AngII AT1 receptor (AT1-R) and NE α2C-adrenergic receptor (α2C-AR) could underlie their functional cross-talk to control NE secretion. Multiple bioluminescence resonance energy transfer and protein complementation assays allowed us to accurately probe the structures and functions of the α2C-AR-AT1-R dimer promoted by ligand binding to individual protomers. We found that dual agonist occupancy resulted in a conformation of the heterodimer different from that induced by active individual protomers and triggered atypical Gs-cAMP-PKA signaling. This specific pharmacological signaling unit was identified in vivo to promote not only NE hypersecretion in sympathetic neurons but also sympathetic hyperactivity in mice. Thus, we uncovered a new process by which GPCR heterodimerization creates an original functional pharmacological entity and that could constitute a promising new target in cardiovascular therapeutics.

Dosage-dependent regulation of cell proliferation and adhesion through dual ß2-adrenergic receptor/cAMP signals.

The role of ß-adrenergic receptors (ß-ARs) remains controversial in normal and tumor breast. Herein we explore the cAMP signaling involved in ß-AR-dependent control of proliferation and adhesion of nontumor human breast cell line MCF-10A. Low concentrations of a ß-agonist, isoproterenol (ISO), promote cell adhesion (87.5% cells remaining adherent to the plastic dishes following specific detachment vs. 35.0% in control, P<0.001), while increasing concentrations further engages an additional 36% inhibition of Erk1/2 phosphorylation (p-Erk1/2)-dependent cell proliferation (P<0.01). Isoproterenol dose response on cell adhesion was fitted to a 2-site curve (EC50(1): 16.5±11.5 fM, EC50(2): 4.08±3.09 nM), while ISO significantly inhibited p-Erk1/2 according to a 1-site model (EC50: 0.25±0.13 nM). Using ß-AR-selective agonist/antagonists and cAMP analogs/inhibitors, we identified a dosage-dependent signaling in which low ISO concentrations target a ß2-AR population localized in raft microdomains and stimulate a Gs/cAMP/Epac/adhesion-signaling module, while higher concentrations engage a concomitant activation of another ß2-AR population outside rafts and inhibit the proliferation by a Gs/cAMP/PKA-dependent signaling module. Our data provide a new molecular basis for the dose-dependent switch of ß-AR signaling. This study also sheds light on a new cAMP pathway core mechanism with a single receptor triggering dual cAMP signaling controlled by PKA or Epac but with different cellular outputs.

Atomic force and electron microscopic-based study of sarcolemmal surface of living cardiomyocytes unveils unexpected mitochondrial shift in heart failure.

Loss of T-tubules (TT), sarcolemmal invaginations of cardiomyocytes (CMs), was recently identified as a general heart failure (HF) hallmark. However, whether TT per se or the overall sarcolemma is altered during HF process is still unknown. In this study, we directly examined sarcolemmal surface topography and physical properties using Atomic Force Microscopy (AFM) in living CMs from healthy and failing mice hearts. We confirmed the presence of highly organized crests and hollows along myofilaments in isolated healthy CMs. Sarcolemma topography was tightly correlated with elasticity, with crests stiffer than hollows and related to the presence of few packed subsarcolemmal mitochondria (SSM) as evidenced by electron microscopy. Three days after myocardial infarction (MI), CMs already exhibit an overall sarcolemma disorganization with general loss of crests topography thus becoming smooth and correlating with a decreased elasticity while interfibrillar mitochondria (IFM), myofilaments alignment and TT network were unaltered. End-stage post-ischemic condition (15days post-MI) exacerbates overall sarcolemma disorganization with, in addition to general loss of crest/hollow periodicity, a significant increase of cell surface stiffness. Strikingly, electron microscopy revealed the total depletion of SSM while some IFM heaps could be visualized beneath the membrane. Accordingly, mitochondrial Ca(2+) studies showed a heterogeneous pattern between SSM and IFM in healthy CMs which disappeared in HF. In vitro, formamide-induced sarcolemmal stress on healthy CMs phenocopied post-ischemic kinetics abnormalities and revealed initial SSM death and crest/hollow disorganization followed by IFM later disarray which moved toward the cell surface and structured heaps correlating with TT loss. This study demonstrates that the loss of crest/hollow organization of CM surface in HF occurs early and precedes disruption of the TT network. It also highlights a general stiffness increased of the CM surface most likely related to atypical IFM heaps while SSM died during HF process. Overall, these results indicate that initial sarcolemmal stress leading to SSM death could underlie subsequent TT disarray and HF setting.

Deciphering biased-agonism complexity reveals a new active AT1 receptor entity.

Functional selectivity of G protein-coupled receptor (GPCR) ligands toward different downstream signals has recently emerged as a general hallmark of this receptor class. However, pleiotropic and crosstalk signaling of GPCRs makes functional selectivity difficult to decode. To look from the initial active receptor point of view, we developed new, highly sensitive and direct bioluminescence resonance energy transfer-based G protein activation probes specific for all G protein isoforms, and we used them to evaluate the G protein-coupling activity of [(1)Sar(4)Ile(8)Ile]-angiotensin II (SII), previously described as an angiotensin II type 1 (AT(1)) receptor-biased agonist that is G protein independent but ß-arrestin selective. By multiplexing assays sensing sequential signaling events, from receptor conformations to downstream signaling, we decoded SII as an agonist stabilizing a G protein-dependent AT(1A) receptor signaling module different from that of the physiological agonist angiotensin II, both in recombinant and primary cells. Thus, a biased agonist does not necessarily select effects from the physiological agonist but may instead stabilize and create a new distinct active pharmacological receptor entity.

Ephrin-B1 is a novel specific component of the lateral membrane of the cardiomyocyte and is essential for the stability of cardiac tissue architecture cohesion.

RATIONALE: Cardiac tissue cohesion relying on highly ordered cardiomyocytes (CM) interactions is critical because most cardiomyopathies are associated with tissue remodeling and architecture alterations. OBJECTIVE: Eph/ephrin system constitutes a ubiquitous system coordinating cellular communications which recently emerged as a major regulator in adult organs. We examined if eph/ephrin could participate in cardiac tissue cyto-organization. METHODS AND RESULTS: We reported the expression of cardiac ephrin-B1 in both endothelial cells and for the first time in CMs where ephrin-B1 localized specifically at the lateral membrane. Ephrin-B1 knock-out (KO) mice progressively developed cardiac tissue disorganization with loss of adult CM rod-shape and sarcomeric and intercalated disk structural disorganization confirmed in CM-specific ephrin-B1 KO mice. CMs lateral membrane exhibited abnormal structure by electron microscopy and notably increased stiffness by atomic force microscopy. In wild-type CMs, ephrin-B1 interacted with claudin-5/ZO-1 complex at the lateral membrane, whereas the complex disappeared in KO/CM-specific ephrin-B1 KO mice. Ephrin-B1 deficiency resulted in decreased mRNA expression of CM basement membrane components and disorganized fibrillar collagen matrix, independently of classical integrin/dystroglycan system. KO/CM-specific ephrin-B1 KO mice exhibited increased left ventricle diameter and delayed atrioventricular conduction. Under pressure overload stress, KO mice were prone to death and exhibited striking tissue disorganization. Finally, failing CMs displayed downregulated ephrin-B1/claudin-5 gene expression linearly related to the ejection fraction. CONCLUSIONS: Ephrin-B1 is necessary for cardiac tissue architecture cohesion by stabilizing the adult CM morphology through regulation of its lateral membrane. Because decreased ephrin-B1 is associated with molecular/functional cardiac defects, it could represent a new actor in the transition toward heart failure.

Ephrin-B1 regulates the adult diastolic function through a late postnatal maturation of cardiomyocyte surface crests.

The rod-shaped adult cardiomyocyte (CM) harbors a unique architecture of its lateral surface with periodic crests, relying on the presence of subsarcolemmal mitochondria (SSM) with unknown role. Here, we investigated the development and functional role of CM crests during the postnatal period. We found in rodents that CM crest maturation occurs late between postnatal day 20 (P20) and P60 through both SSM biogenesis, swelling and crest-crest lateral interactions between adjacent CM, promoting tissue compaction. At the functional level, we showed that the P20-P60 period is dedicated to the improvement of relaxation. Interestingly, crest maturation specifically contributes to an atypical CM hypertrophy of its short axis, without myofibril addition, but relying on CM lateral stretching. Mechanistically, using constitutive and conditional CM-specific knock-out mice, we identified ephrin-B1, a lateral membrane stabilizer, as a molecular determinant of P20-P60 crest maturation, governing both the CM lateral stretch and the diastolic function, thus highly suggesting a link between crest maturity and diastole. Remarkably, while young adult CM-specific Efnb1 KO mice essentially exhibit an impairment of the ventricular diastole with preserved ejection fraction and exercise intolerance, they progressively switch toward systolic heart failure with 100% KO mice dying after 13 months, indicative of a critical role of CM-ephrin-B1 in the adult heart function. This study highlights the molecular determinants and the biological implication of a new late P20-P60 postnatal developmental stage of the heart in rodents during which, in part, ephrin-B1 specifically regulates the maturation of the CM surface crests and of the diastolic function.

[Adult resident cardiomyocytes wake up: new axis for cardiac tissue regeneration]. / Le réveil des cardiomyocytes adultes résidents - Une nouvelle piste de régénération cardiaque ?

All cardiomyopathies and more specifically myocardial infarction always evolve to cardiomyocytes death and the ensuing heart failure setting. So far, cardiac regenerative medicine has focused on the use of stem cells and completely ignored the resident cardiomyocytes, assumed in a postmitotic state. However, recent findings in zebrafish and mammalians challenge this view and suggest that these cells have some capacity to proliferate and can contribute to heart regeneration. In this review, we propose an overall synthesis about knowledge of the proliferative and regenerative capacities of resident cardiomyocytes, dealing with some mechanistic aspects. In the future, the accurate identification of molecular mechanisms allowing wake-up of resident cardiomyocyte proliferation will certainly open new therapeutic avenues in cardiac regeneration.

Age and Gender Differences in Cardiovascular Autonomic Failure in the Transgenic PLP-syn Mouse, a Model of Multiple System Atrophy.

Multiple system atrophy (MSA) is a rare and progressive neurodegenerative disorder. Autonomic failure (AF) is one main clinical feature which has a significant impact on health-related quality of life. The neuropathological hallmark of MSA is the abnormal accumulation of α-synuclein in oligodendrocytes forming glial cytoplasmic inclusions. Only little is known about gender and age differences in AF in MSA. This study was carried out in 6 and 12 months old transgenic PLP-α-syn and WT male and female mice. Heart rate variability (HRV) was assessed both in time, frequential and non-linear domains. Baroreflex sensitivity (BRS) was estimated by the sequence method. Duration of ventricular depolarization and repolarization (QT/QTc intervals) were evaluated from the ECG signals. Three-way ANOVA (genotype x gender x age) with Sidak's method post-hoc was used to analyze data. BRS was significantly changed in PLP-α-syn mice and was age-dependent. QT and QTc intervals were not significantly modified in PLP-α-syn mice. An impaired HRV was observed at 12 months of age in PLP-α-syn female but not in male mice, indicative of cardiovascular AF.

Atomic force microscopy-single-molecule force spectroscopy unveils GPCR cell surface architecture.

G protein-coupled receptors (GPCRs) form the largest family of cell surface receptors. Despite considerable insights into their pharmacology, the GPCR architecture at the cell surface still remains largely unexplored. Herein, we present the specific unfolding of different GPCRs at the surface of living mammalian cells by atomic force microscopy-based single molecule force spectroscopy (AFM-SMFS). Mathematical analysis of the GPCR unfolding distances at resting state revealed the presence of different receptor populations relying on distinct oligomeric states which are receptor-specific and receptor expression-dependent. Moreover, we show that the oligomer size dictates the receptor spatial organization with nanoclusters of high-order oligomers while lower-order complexes spread over the whole cell surface. Finally, the receptor activity reshapes both the oligomeric populations and their spatial arrangement. These results add an additional level of complexity to the GPCR pharmacology until now considered to arise from a single receptor population at the cell surface.

How does head position induced intracranial pressure changes impact sympathetic activity and cerebral blood flow?

PURPOSE: Acute head-down-tilt (HDT) simulates short duration hemodynamic impact of microgravity. We sought to determine whether an increase in ICP caused by acute HDT affects sympathetic nervous system activity and cerebral blood flow velocities (CBFV) in healthy male volunteers. METHODS: HDT protocol was established as follows: basal condition immediately followed by gradual negative angles (-10°, -20° and -30°) lasting 10mn and then a return to basal condition. Velocities in the MCA (CBFV) were monitored using TCD. Sympathetic activity was assessed using MSNA. Baroreflex sensitivity (BRS) was measured using the sequence method. ICP changes were assessed using ultrasonography of the optic nerve sheath diameter (ONSD). Cerebral autoregulation (CA) was evaluated by transfer function and the autoregulatory index (Mxa). RESULTS: Twelve male volunteers (age: 35 ± 2 years) were included. Neither blood pressure nor heart rate was significantly modified during HDT. ONSD increased significantly at each step of HDT and remained elevated during Recovery. MSNA burst incidence increased at -30°. A positive correlation between variations in ONSD and variations in MSNA burst incidence was observed at -20°. CBFV were significantly diminished at -20° and -30. In the LF band, the transfer function coherence was reduced at -30° and the transfer function phase was increased at -30° and during Recovery. DISCUSSION: We found that an acute though modest increase in ICP induced by HDT was associated with an increase of sympathetic activity as assessed by MSNA, and with a reduction of CBFV with preserved CA.