Key points
Cerebellar Purkinje cells (PCs) generate two types of action potentials, simple and complex spikes. Although they are generated by distinct mechanisms, interactions between the two spike ...types exist.
Zebrin staining produces alternating positive and negative stripes of PCs across most of the cerebellar cortex. Thus, here we compared simple spike–complex spike interactions both within and across zebrin populations.
Simple spike activity undergoes a complex modulation preceding and following a complex spike. The amplitudes of the pre‐ and post‐complex spike modulation phases were correlated across PCs. On average, the modulation was larger for PCs in zebrin positive regions.
Correlations between aspects of the complex spike waveform and simple spike activity were found, some of which varied between zebrin positive and negative PCs.
The implications of the results are discussed with regard to hypotheses that complex spikes are triggered by rises in simple spike activity for either motor learning or homeostatic functions.
Purkinje cells (PCs) generate two types of action potentials, called simple and complex spikes (SSs and CSs). We first investigated the CS‐associated modulation of SS activity and its relationship to the zebrin status of the PC. The modulation pattern consisted of a pre‐CS rise in SS activity, and then, following the CS, a pause, a rebound, and finally a late inhibition of SS activity for both zebrin positive (Z+) and negative (Z−) cells, though the amplitudes of the phases were larger in Z+ cells. Moreover, the amplitudes of the pre‐CS rise with the late inhibitory phase of the modulation were correlated across PCs. In contrast, correlations between modulation phases across CSs of individual PCs were generally weak. Next, the relationship between CS spikelets and SS activity was investigated. The number of spikelets/CS correlated with the average SS firing rate only for Z+ cells. In contrast, correlations across CSs between spikelet numbers and the amplitudes of the SS modulation phases were generally weak. Division of spikelets into likely axonally propagated and non‐propagated groups (based on their interspikelet interval) showed that the correlation of spikelet number with SS firing rate primarily reflected a relationship with non‐propagated spikelets. In sum, the results show both zebrin‐related and non‐zebrin‐related physiological heterogeneity in SS–CS interactions among PCs, which suggests that the cerebellar cortex is more functionally diverse than is assumed by standard theories of cerebellar function.
Key points
Cerebellar Purkinje cells (PCs) generate two types of action potentials, simple and complex spikes. Although they are generated by distinct mechanisms, interactions between the two spike types exist.
Zebrin staining produces alternating positive and negative stripes of PCs across most of the cerebellar cortex. Thus, here we compared simple spike–complex spike interactions both within and across zebrin populations.
Simple spike activity undergoes a complex modulation preceding and following a complex spike. The amplitudes of the pre‐ and post‐complex spike modulation phases were correlated across PCs. On average, the modulation was larger for PCs in zebrin positive regions.
Correlations between aspects of the complex spike waveform and simple spike activity were found, some of which varied between zebrin positive and negative PCs.
The implications of the results are discussed with regard to hypotheses that complex spikes are triggered by rises in simple spike activity for either motor learning or homeostatic functions.
Aldosterone-producing adenoma and bilateral adrenal hyperplasia account for >90% of all primary aldosteronism cases. Distinguishing between bilateral and unilateral disease is of fundamental ...importance because it allows targeted therapy. Adrenal vein sampling (AVS) is the only reliable means to preoperatively differentiate between unilateral and bilateral subtypes. A rare but serious complication of AVS is an adrenal hemorrhage (AH). We retrospectively examined in detail 24 cases of AH during AVS in 6 different referral hypertension centers. AH more often affected the right adrenal (n=18) than the left (n=5, P<0.001); 1 bilateral. Median duration of experience of the radiologist in AVS at the time of AH was 5.0 years (0.6-7.8) and AH occurred with both highly experienced (>10 years) and less experienced radiologists. Of 9 patients who suffered AH in the gland contralateral to an aldosterone-producing adenoma and who underwent complete (n=6) or partial (n=3) unilateral adrenalectomy, only one required long-term corticosteroid replacement for adrenal insufficiency. No reduction in blood pressure or biochemical resolution of primary aldosteronism occurred in any of those patients who experienced AH in the gland ipsilateral to an aldosterone-producing adenoma (n=6) or who had bilateral adrenal hyperplasia (n=9). No patient required invasive treatments to control bleeding or blood transfusion. In conclusion, AH usually has a positive outcome causing either no or minor effects on adrenal function, and AVS should remain the best approach to primary aldosteronism subtype differentiation.
Although monocytes/macrophages are considered important in atherogenesis, their role in established plaques is unclear. For example, macrophage content is associated with plaque instability, but ...their loss through cell death is observed at sites of plaque rupture. To examine the role of monocytes/macrophages in atherosclerosis, we developed CD11b–diphtheria toxin (DT) receptor (DTR) transgenic mice, whereby administration of DT selectively kills monocytes/macrophages. DT treatment reduced peripheral blood monocytes and tissue macrophages and inhibited macrophage function in CD11b-DTR mice and apolipoprotein E–null (apoE) mice transplanted with CD11b-DTR bone marrow. In atherogenesis experiments, DT markedly reduced plaque development and altered plaque composition, reducing collagen content and necrotic core formation. In mice with established plaques, acute DT treatment induced macrophage apoptosis and reduced macrophage content but did not induce plaque inflammation, thrombosis, or rupture. Furthermore, despite a 50% reduction in monocytes, chronic DT treatment of these mice did not alter plaque extent or composition, most likely because of ongoing recruitment/proliferation of monocytes with recovery of macrophage content. We conclude that monocytes/macrophages are critical to atherogenesis, but established plaques are more resistant to reductions in monocytes.
To assess cardiovascular magnetic resonance (CMR) measured myocardial perfusion reserve (MPR) and exercise testing in asymptomatic patients with moderate-severe AS.
Multi-centre, prospective, ...observational study, with blinded analysis of CMR data. Patients underwent adenosine stress CMR, symptom-limited exercise testing (ETT) and echocardiography and were followed up for 12-30 months. The primary outcome was a composite of: typical AS symptoms necessitating referral for AVR, cardiovascular death and major adverse cardiovascular events. 174 patients were recruited: mean age 66.2 ± 13.34 years, 76% male, peak velocity 3.86 ± 0.56 m/s and aortic valve area index 0.57 ± 0.14 cm2/m2. A primary outcome occurred in 47 (27%) patients over a median follow-up of 374 (IQR 351-498) days. The mean MPR in those with and without a primary outcome was 2.06 ± 0.65 and 2.34 ± 0.70 (P = 0.022), while the incidence of a symptom-limited ETT was 45.7% and 27.0% (P = 0.020), respectively. MPR showed moderate association with outcome area under curve (AUC) = 0.61 (0.52-0.71, P = 0.020), as did exercise testing (AUC = 0.59 (0.51-0.68, P = 0.027), with no significant difference between the two.
MPR was associated with symptom-onset in initially asymptomatic patients with AS, but with moderate accuracy and was not superior to symptom-limited exercise testing. ClinicalTrials.gov (NCT01658345).
Food allergy is a major health problem affecting 5% to 10% of the population in developed nations, including an estimated 32 million Americans. Despite the large number of patients suffering from ...food allergies, up until the end of January 2020, no treatment for food allergies had been approved by the US Food and Drug Administration. The only options were avoidance of food allergen triggers and acute management of allergic reactions. A considerable body of data exists supporting oral immunotherapy (OIT) as a promising, novel treatment option, including that for the now Food and Drug Administration–approved peanut OIT product Palforzia (Aimmune Therapeutics, Brisbane, Calif). However, data for long-term quality-of-life improvement with OIT varies, depending on the measures used for analysis. Like many therapies, OIT is not without potential harms, and burdens, and the evaluation of patient-specific risk-benefit ratio of food OIT produces challenges for clinicians and patients alike, with many unanswered questions. Food Allergy Research & Education organized the Oral Immunotherapy for Food Allergy Summit on November 6, 2019, modeled after the PRACTALL sessions between the European Academy of Allergy and Clinical Immunology and the American Academy of Allergy, Asthma & Immunology to address these critical issues. Health care providers, patient representatives, researchers, regulators, and food allergy advocates came together to discuss OIT and identify areas of common ground as well as gaps in existing research and areas of uncertainty and disagreement. The purpose of this article was to summarize that discussion and facilitate collaboration among clinicians and patients to help them make better-informed decisions about offering and accepting OIT, respectively, as a therapeutic option.
Typical and atypical smooth muscle cells (TSMCs and ASMCs, respectively) and interstitial cells (ICs) within the pacemaker region of the mouse renal pelvis were examined using focused ion beam ...scanning electron (FIB SEM) tomography, immunohistochemistry and Ca
2+
imaging. Individual cells within 500–900 electron micrograph stacks were volume rendered and associations with their neighbours established. ‘Ribbon-shaped’, Ano1 Cl
−
channel immuno-reactive ICs were present in the adventitia and the sub-urothelial space adjacent to the TSMC layer. ICs in the proximal renal pelvis were immuno-reactive to antibodies for Ca
V
3.1 and hyperpolarization-activated cation nucleotide-gated isoform 3 (HCN3) channel sub-units, while basal-epithelial cells (BECs) were intensely immuno-reactive to Kv7.5 channel antibodies. Adventitial to the TSMC layer, ASMCs formed close appositions with TSMCs and ICs. The T-type Ca
2+
channel blocker, Ni
2+
(10–200 μM), reduced the frequency while the L-type Ca
2+
channel blocker (1 μM nifedipine) reduced the amplitude of propagating Ca
2+
waves and contractions in the TSMC layer. Upon complete suppression of Ca
2+
entry through TSMC Ca
2+
channels, ASMCs displayed high-frequency (6 min
−1
) Ca
2+
transients, and ICs distributed into two populations of cells firing at 1 and 3 min
−1
, respectively. IC Ca
2+
transients periodically (every 3–5 min
−1
) summed into bursts which doubled the frequency of ASMC Ca
2+
transient firing. Synchronized IC bursting and the acceleration of ASMC firing were inhibited upon blockade of HCN channels with ZD7288 or cell-to-cell coupling with carbenoxolone. While ASMCs appear to be the primary pacemaker driving pyeloureteric peristalsis, it was concluded that sub-urothelial HCN3(+), Ca
V
3.1(+) ICs can accelerate ASMC Ca
2+
signalling.
In human cells, approximately 30% of all polypeptides enter the secretory pathway at the level of the endoplasmic reticulum (ER). This process involves cleavable amino-terminal signal peptides (SPs) ...or more or less amino-terminal transmembrane helices (TMHs), which serve as targeting determinants, at the level of the precursor polypeptides and a multitude of cytosolic and ER proteins, which facilitate their ER import. Alone or in combination SPs and TMHs guarantee the initial ER targeting as well as the subsequent membrane integration or translocation. Cytosolic SRP and SR, its receptor in the ER membrane, mediate cotranslational targeting of most nascent precursor polypeptide chains to the polypeptide-conducting Sec61 complex in the ER membrane. Alternatively, fully-synthesized precursor polypeptides and certain nascent precursor polypeptides are targeted to the ER membrane by either the PEX-, SND-, or TRC-pathway. Although these targeting pathways may have overlapping functions, the question arises how relevant this is under cellular conditions and which features of SPs and precursor polypeptides determine preference for a certain pathway. Irrespective of their targeting pathway(s), most precursor polypeptides are integrated into or translocated across the ER membrane via the Sec61 channel. For some precursor polypeptides specific Sec61 interaction partners have to support the gating of the channel to the open state, again raising the question why and when this is the case. Recent progress shed light on the client spectrum and specificities of some auxiliary components, including Sec62/Sec63, TRAM1 protein, and TRAP. To address the question which precursors use a certain pathway or component in intact human cells, i.e., under conditions of fast translation rates and molecular crowding, in the presence of competing precursors, different targeting organelles, and relevant stoichiometries of the involved components, siRNA-mediated depletion of single targeting or transport components in HeLa cells was combined with label-free quantitative proteomics and differential protein abundance analysis. Here, we present a summary of the experimental approach as well as the resulting differential protein abundance analyses and discuss their mechanistic implications in light of the available structural data.
Abstract Background and purpose To analyse dose–response relationships for local control of cervical cancer after MR image-guided brachytherapy (IGBT) based on dose–volume histogram parameters. ...Methods and materials The analysis includes 141 patients with cervix cancer (stages IB–IVA) treated with 45–50.4 Gy EBRT ± cisplatin plus 4 × 7 Gy IGBT. Gross tumour volume (GTV), high risk clinical target volume (HR CTV) and intermediate risk CTV (IR CTV) were delineated and DVH parameters (D90, D100) were assessed. Doses were converted to the equivalent dose in 2 Gy (EQD2) using linear-quadratic model ( α / β = 10 Gy). Groups of patients were formed according to tumour size at diagnosis (GTVD ) of 2–5 cm (group 1) or >5 cm (2), with subgroups of the latter for HR CTV size at first IGBT 2–5 cm (2a) or >5 cm (2b). Dose–response dependence for local recurrence was evaluated by logit analysis. Results Eighteen local recurrences in the true pelvis were observed. Dose–response analyses revealed a significant effect of HR CTV D100 ( p = 0.02) and D90 ( p = 0.005). The ED50-values for tumour control were 33 ± 15 Gy (D100) and 45 ± 19 Gy (D90). ED90-values were 67 Gy (95% confidence interval 50;104) and 86 Gy 77;113, respectively. Conclusions A significant dependence of local control on D100 and D90 for HR CTV was found. Tumour control rates of >90% can be expected at doses >67 Gy and 86 Gy, respectively.
In mammalian cells, proteins are typically translocated across the endoplasmic reticulum (ER) membrane in a co-translational mode by the ER protein translocon, comprising the protein-conducting ...channel Sec61 and additional complexes involved in nascent chain processing and translocation. As an integral component of the translocon, the oligosaccharyl-transferase complex (OST) catalyses co-translational N-glycosylation, one of the most common protein modifications in eukaryotic cells. Here we use cryoelectron tomography, cryoelectron microscopy single-particle analysis and small interfering RNA-mediated gene silencing to determine the overall structure, oligomeric state and position of OST in the native ER protein translocon of mammalian cells in unprecedented detail. The observed positioning of OST in close proximity to Sec61 provides a basis for understanding how protein translocation into the ER and glycosylation of nascent proteins are structurally coupled. The overall spatial organization of the native translocon, as determined here, serves as a reliable framework for further hypothesis-driven studies.
After repair of congenital heart defects, stress responses in the pulmonary circulation of fourteen infants produced by a brief endotracheal suctioning procedure were studied before and after ...fentanyl (25 micrograms/kg). The total time of disconnection from the ventilator with suctioning (FIO2 1.0) was limited to no more than 15 sec to avoid alveolar hypoxia. Before fentanyl, marked increases occurred in mean pulmonary artery pressure and pulmonary vascular resistance index with suctioning, whereas only mild increases in heart rate and mean systemic arterial pressure occurred. All of these increases with suctioning were almost completely abolished by 25 micrograms/kg fentanyl. We conclude that suctioning or other broncho-carinal stimulation can produce a marked pulmonary vasoconstrictive response in infants, which is blunted by fentanyl. This response is separate from that produced by hypoxic pulmonary vasoconstriction associated with prolonged clinical suctioning procedures or with loss of airway.
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