Myogenic contractions of a somatic muscle powers rhythmic flow of hemolymph through Drosophila antennae and generates brain pulsations

Hemolymph is driven through the antennae of Drosophila melanogaster by the rhythmic contraction of muscle 16 (m16), which runs through the brain. Contraction of m16 results in the expansion of an elastic ampulla, opening ostia and filling the ampulla. Relaxation of the ampullary membrane forces hemolymph through vessels into the antennae. We show that m16 is an auto-active rhythmic somatic muscle. The activity of m16 leads to the rapid perfusion of the antenna by hemolymph. In addition, it leads to the rhythmic agitation of the brain, which could be important for clearing the interstitial space.

Association of intracranial arteriovenous malformation embolization with more rapid rate of perfusion in the peri-nidal region on color-coded quantitative digital subtraction angiography

BackgroundHemodynamic alterations post-embolization of intracranial arteriovenous malformations (AVMs) may cause delayed edema/hemorrhage in brain parenchyma adjacent to the lesion.ObjectiveTo quantify and compare cerebral perfusion changes in the peri-AVM territory pre- and post-embolization using color-coded quantitative digital subtraction angiography (q-DSA).MethodsPediatric intracranial AVM embolization procedures performed over a 5 year period were included. DSA images of all patients were retrospectively assessed using syngo iFlow. Regions of interest (ROI) were selected on anteroposterior and lateral q-DSA views: three in the peri-AVM region; two in parenchyma distant from the AVM. Time-to-peak (TTP) contrast enhancement of ROIs and ∆TTP (TTP at the selected ROI minus TTP at either the ipsilateral internal carotid/vertebral artery) were measured.Result19 pediatric patients with 19 AVMs (9 males/10 females, mean age 12 years) underwent intracranial AVM embolization: 15/19 AVMs were supplied by the anterior circulation and 4/19 by the posterior circulation. Blood flow was significantly slower post-embolization in the draining vein (19/19) (p<0.01), and the venous sinus outflow (17/19) (p<0.01), by mean difference of 2.01±1.31 s and 1.74±2.04 s. There was significantly increased peri-AVM parenchymal perfusion post-embolization (∆TTP=2.20±0.48 s) compared with pre-embolization (∆TTP=2.52±0.42 s), by an average ∆TTP of 0.33±0.53 s (p=0.014). In contrast, there was no perfusion difference (∆TTP=0.03±0.20 s, p=0.8) between pre- and post-embolization in the distant parenchyma. The size of the AVM was not correlated with change in peri-nidal parenchymal perfusion (r=−0.136, p=0.579).ConclusionThis study demonstrates more rapid perfusion in the peri-nidal brain parenchyma post-embolization of the AVM, which supports the theory that increased perfusion in normal tissue surrounding the AVM after embolization may underlie some post-procedural complications.

Abstract WP304: Real-World Experience with the RAPID Platform to Screen Acute Ischemic Stroke Patients at a U.S. Comprehensive Stroke Center

Introduction: Mechanical thrombectomy (MT) is an important part of acute ischemic stroke (AIS) treatment. Recent trials of MT beyond the 6-hour window have utilized RAPID perfusion imaging for patient selection. The utility of this method is established in patients with large vessel occlusions (LVO) but screening efficiency in real-world practice remains unknown. We present the experience of a single, large volume, Comprehensive Stroke Center (CSC) utilizing RAPID to screen patients for LVO and MT. Methods: We performed a retrospective analysis of prospectively collected consecutive patients who presented to our emergency department (ED) between 01/2018 to 06/2019 with suspected LVO. Protocol was based on 2018 AHA guideline Level IA recommendations and followed DAWN and DEFUSE-3 time and imaging parameters. Patients who underwent RAPID imaging were selected for inclusion. Results: 865 patients met criteria for RAPID perfusion imaging (median age 67, females 52%, outside hospital transfers 29%). Of these, 178 (21% of total) were confirmed to have an LVO (40% ED presentation, 10% inpatient, 50% transfer). For patients presenting to the ED (N=509), 14% had an LVO (median NIHSS 13 [IQR 8-19]), of which 41% underwent MT. Mean CTP core and penumbra volume was 25mL and 100mL respectively. Number needed-to-screen in the ED cohort was 7 to detect LVO and 17 to perform MT. Transfer patients showed no significant difference in LVO detection or MT rates compared to ED patients (56%, p=0.3). Conclusions: In ED-presenting patients at a CSC, the number of RAPID perfusion imaging studies needed to detect an additional case of LVO was 7.1, and to perform an additional MT was 17.4. Current AHA Class IA recommendations for evaluation and treatment of AIS yield a reasonably high rate of LVO detection and subsequent MT in real-world practice. Additional multicenter data will be useful to establish benchmarks and improve screening efficiency.

Unitary Ca2+ current through recombinant type 3 InsP3 receptor channels under physiological ionic conditions

The ubiquitous inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) channel, localized primarily in the endoplasmic reticulum (ER) membrane, releases Ca2+ into the cytoplasm upon binding InsP3, generating and modulating intracellular Ca2+ signals that regulate numerous physiological processes. Together with the number of channels activated and the open probability of the active channels, the size of the unitary Ca2+ current (iCa) passing through an open InsP3R channel determines the amount of Ca2+ released from the ER store, and thus the amplitude and the spatial and temporal nature of Ca2+ signals generated in response to extracellular stimuli. Despite its significance, iCa for InsP3R channels in physiological ionic conditions has not been directly measured. Here, we report the first measurement of iCa through an InsP3R channel in its native membrane environment under physiological ionic conditions. Nuclear patch clamp electrophysiology with rapid perfusion solution exchanges was used to study the conductance properties of recombinant homotetrameric rat type 3 InsP3R channels. Within physiological ranges of free Ca2+ concentrations in the ER lumen ([Ca2+]ER), free cytoplasmic [Ca2+] ([Ca2+]i), and symmetric free [Mg2+] ([Mg2+]f), the iCa–[Ca2+]ER relation was linear, with no detectable dependence on [Mg2+]f. iCa was 0.15 ± 0.01 pA for a filled ER store with 500 µM [Ca2+]ER. The iCa–[Ca2+]ER relation suggests that Ca2+ released by an InsP3R channel raises [Ca2+]i near the open channel to ∼13–70 µM, depending on [Ca2+]ER. These measurements have implications for the activities of nearby InsP3-liganded InsP3R channels, and they confirm that Ca2+ released by an open InsP3R channel is sufficient to activate neighboring channels at appropriate distances away, promoting Ca2+-induced Ca2+ release.

Pentobarbital Produces Activation and Block of α1β2γ2S GABAA Receptors in Rapidly Perfused Whole Cells and Membrane Patches: Divergent Results Can Be Explained by Pharmacokinetics

Millimolar concentrations of the barbiturate pentobarbital (PB) activate γ-aminobutyric acid (GABA) type A receptors (GABARs) and cause blockade reported by a paradoxical current increase or “tail” upon washout. To explore the mechanism of blockade, we investigated PB-triggered currents of recombinant α1β2γ2S GABARs in whole cells and outside-out membrane patches using rapid perfusion. Whole cell currents showed characteristic bell-shaped concentration dependence where high concentrations triggered tail currents with peak amplitudes similar to those during PB application. Tail current time courses could not be described by multi-exponential functions at high concentrations (≥3,000 μM). Deactivation time course decayed over seconds and was slowed by increasing PB concentration and application time. In contrast, macropatch tail currents manifested eightfold greater relative amplitude, were described by multi-exponential functions, and had millisecond rise times; deactivation occurred over fractions of seconds and was insensitive to PB concentration and application time. A parsimonious gating model was constructed that accounts for macropatch results (“patch” model). Lipophilic drug molecules migrate slowly through cells due to avid partitioning into lipophilic subcellular compartments. Inclusion of such a pharmacokinetic compartment into the patch model introduced a slow kinetic component in the extracellular exchange time course, thereby providing recapitulation of divergent whole cell results. GABA co-application potentiated PB blockade. Overall, the results indicate that block is produced by PB concentrations sixfold lower than for activation involving at least three inhibitory PB binding sites, suggest a role of blocked channels in GABA-triggered activity at therapeutic PB concentrations, and raise an important technical question regarding the effective rate of exchange during rapid perfusion of whole cells with PB.

Planar Phospholipid Membrane Formation in Open Well Thermoplastic Chips

A key requirement for the effective study of interactions between analytes and ion channels is the ability to dynamically vary analyte type and concentration to a membrane-bound ion channel within a planar phospholipid membrane (PPM). Here an open well microfluidic PPM apparatus supporting dynamic perfusion is presented. The plastic chip supports the manual formation of bilayer membranes that are resistant to pressure disturbances during perfusion with stability on the order of several hours. Using a chamber volume of 20 μL and a flow rate of 0.5 μL/min, the system enables rapid perfusion without breaking the membrane. The perfusion capability is demonstrated through gramicidin ion channel measurements.