A Position Statement on the Utility of Interval Imaging in Standard of Care Brain Tumour Management: Defining the Evidence Gap and Opportunities for Future Research.

OBJECTIV E: To summarise current evidence for the utility of interval imaging in monitoring disease in adult brain tumours, and to develop a position for future evidence gathering while incorporating the application of data science and health economics. METHODS: Experts in 'interval imaging' (imaging at pre-planned time-points to assess tumour status); data science; health economics, trial management of adult brain tumours, and patient representatives convened in London, UK. The current evidence on the use of interval imaging for monitoring brain tumours was reviewed. To improve the evidence that interval imaging has a role in disease management, we discussed specific themes of data science, health economics, statistical considerations, patient and carer perspectives, and multi-centre study design. Suggestions for future studies aimed at filling knowledge gaps were discussed. RESULTS: Meningioma and glioma were identified as priorities for interval imaging utility analysis. The "monitoring biomarkers" most commonly used in adult brain tumour patients were standard structural MRI features. Interval imaging was commonly scheduled to provide reported imaging prior to planned, regular clinic visits. There is limited evidence relating interval imaging in the absence of clinical deterioration to management change that alters morbidity, mortality, quality of life, or resource use. Progression-free survival is confounded as an outcome measure when using structural MRI in glioma. Uncertainty from imaging causes distress for some patients and their caregivers, while for others it provides an important indicator of disease activity. Any study design that changes imaging regimens should consider the potential for influencing current or planned therapeutic trials, ensure that opportunity costs are measured, and capture indirect benefits and added value. CONCLUSION: Evidence for the value, and therefore utility, of regular interval imaging is currently lacking. Ongoing collaborative efforts will improve trial design and generate the evidence to optimise monitoring imaging biomarkers in standard of care brain tumour management.

Lack of influence of dexmedetomidine on rat glomus cell response to hypoxia, and on mouse acute hypoxic ventilatory response.

Background and Aims: There is a lack of basic science data on the effect of dexmedetomidine on the hypoxic chemosensory reflex with both depression and stimulation suggested. The primary aim of this study was to assess if dexmedetomidine inhibited the cellular response to hypoxia in rat carotid body glomus cells, the cells of the organs mediating acute hypoxic ventilatory response (AHVR). Additionally, we used a small sample of mice to assess if there was any large influence of subsedative doses of dexmedetomidine on AHVR. Material and Methods: In the primary study, glomus cells isolated from neonatal rats were used to study the effect of 0.1 nM (n = 9) and 1 nM (n = 13) dexmedetomidine on hypoxia-elicited intracellular calcium [Ca2%]i influx using ratiometric fluorimetry. Secondarily, whole animal unrestrained plethysmography was used to study AHVR in a total of 8 age-matched C57BL6 mice, divided on successive days into two groups of four mice randomly assigned to receive sub-sedative doses of 5, 50, or 500 µg.kg-1 dexmedetomidine versus control in a crossover study design (total n = 12 exposures to drug with n = 12 controls). Results: There was no effect of dexmedetomidine on the hypoxia-elicited increase in [Ca2%]i in glomus cells (a mean ± SEM increase of 95 ± 32 nM from baseline with control hypoxia, 124 ± 41 nM with 0.1 nM dexmedetomidine; P = 0.514). In intact mice, dexmedetomidine had no effect on baseline ventilation during air-breathing (4.01 ± 0.3 ml.g-1.min-1 in control and 2.99 ± 0.5 ml.g-1.min-1 with 500 µg.kg-1 dexmedetomidine, the highest dose; P = 0.081) or on AHVR (136 ± 19% increase from baseline in control, 152 ± 46% with 500 µg.kg-1 dexmedetomidine, the highest dose; P = 0.536). Conclusion: Dexmedetomidine had no effect on the cellular responses to hypoxia. We conclude that it unlikely acts via inhibition of oxygen sensing at the glomus cell. The respiratory chemoreflex effects of this drug remain an open question. In our small sample of intact mice, hypoxic chemoreflex responses and basal breathing were preserved.

Competitive Interactions between Halothane and Isoflurane at the Carotid Body and TASK Channels.

BACKGROUND: The degree to which different volatile anesthetics depress carotid body hypoxic response relates to their ability to activate TASK potassium channels. Most commonly, volatile anesthetic pairs act additively at their molecular targets. We examined whether this applied to carotid body TASK channels. METHODS: We studied halothane and isoflurane effects on hypoxia-evoked rise in intracellular calcium (Ca2+i, using the indicator Indo-1) in isolated neonatal rat glomus cells, and TASK single-channel activity (patch clamping) in native glomus cells and HEK293 cell line cells transiently expressing TASK-1. RESULTS: Halothane (5%) depressed glomus cell Ca2+i hypoxic response (mean ± SD, 94 ± 4% depression; P < 0.001 vs. control). Isoflurane (5%) had a less pronounced effect (53 ± 10% depression; P < 0.001 vs. halothane). A mix of 3% isoflurane/1.5% halothane depressed cell Ca2+i response (51 ± 17% depression) to a lesser degree than 1.5% halothane alone (79 ± 15%; P = 0.001), but similar to 3% isoflurane alone (44 ± 22%; P = 0.224), indicating subadditivity. Halothane and isoflurane increased glomus cell TASK-1/TASK-3 activity, but mixes had a lesser effect than that seen with halothane alone: 4% halothane/4% isoflurane yielded channel open probabilities 127 ± 55% above control, versus 226 ± 12% for 4% halothane alone (P = 0.009). Finally, in HEK293 cell line cells, progressively adding isoflurane (1.5 to 5%) to halothane (2.5%) reduced TASK-1 channel activity from 120 ± 38% above control, to 88 ± 48% (P = 0.034). CONCLUSIONS: In all three experimental models, the effects of isoflurane and halothane combinations were quantitatively consistent with the modeling of weak and strong agonists competing at a common receptor on the TASK channel.

Influence of propofol on isolated neonatal rat carotid body glomus cell response to hypoxia and hypercapnia.

In humans the intravenous anaesthetic propofol depresses ventilatory responses to hypoxia and CO2. Animal studies suggest that this may in part be due to inhibition of synaptic transmission between chemoreceptor glomus cells of the carotid body and the afferent carotid sinus nerve. It is however unknown if propofol can also act directly on the glomus cell. Here we report that propofol can indeed inhibit intracellular Ca2+ responses to hypoxia and hypercapnia in isolated rat glomus cells. Neither this propofol effect, nor the glomus cell response to hypoxia in the absence of propofol, were influenced by GABA receptor activation (using GABA, muscimol and baclofen) or inhibition (using bicuculline and 5-aminovaleric acid). Suggesting that these effects of propofol are not mediated through GABA receptors. Propofol inhibited calcium responses to nicotine in glomus cells but the nicotinic antagonists vecuronium and methyllycaconitine did not inhibit calcium responses to hypoxia. TASK channel activity was not altered by propofol. The glomus cell Ca2+ response to depolarisation with 30 mM K+ was however modestly inhibited by propofol. In summary we conclude that propofol does have a direct effect upon hypoxia signalling in isolated type-1 cells and that this may be partially due to its ability to inhibit voltage gated Ca2+v channels. We also note that propofol has the capacity to supress glomus cell excitation via nicotinic receptors and may therefore also interfere with paracrine/autocrine cholinergic signalling in the intact organ. The effects of propofol on chemoreceptor function are however clearly complex and require further investigation.

A1899, PK-THPP, ML365, and Doxapram inhibit endogenous TASK channels and excite calcium signaling in carotid body type-1 cells.

Sensing of hypoxia and acidosis in arterial chemoreceptors is thought to be mediated through the inhibition of TASK and possibly other (e.g., BKCa ) potassium channels which leads to membrane depolarization, voltage-gated Ca-entry, and neurosecretion. Here, we investigate the effects of pharmacological inhibitors on TASK channel activity and [Ca2+ ]i -signaling in isolated neonatal rat type-1 cells. PK-THPP inhibited TASK channel activity in cell attached patches by up to 90% (at 400 nmol/L). A1899 inhibited TASK channel activity by 35% at 400 nmol/L. PK-THPP, A1899 and Ml 365 all evoked a rapid increase in type-1 cell [Ca2+ ]i . These [Ca2+ ]i responses were abolished in Ca2+ -free solution and greatly attenuated by Ni2+ (2 mM) suggesting that depolarization and voltage-gated Ca2+ -entry mediated the rise in [Ca2+ ]i. Doxapram (50 µmol/L), a respiratory stimulant, also inhibited type-1 cell TASK channel activity and increased [Ca2+ ]i. . We also tested the effects of combined inhibition of BKCa and TASK channels. TEA (5 mmol/L) slightly increased [Ca2+ ]i in the presence of PK-THPP and A1899. Paxilline (300 nM) and iberiotoxin (50 nmol/L) also slightly increased [Ca2+ ]i in the presence of A1899 but not in the presence of PK-THPP. In general [Ca2+ ]i responses to TASK inhibitors, alone or in combination with BKCa inhibitors, were smaller than the [Ca2+ ]i responses evoked by hypoxia. These data confirm that TASK channel inhibition is capable of evoking membrane depolarization and robust voltage-gated Ca2+ -entry but suggest that this, even with concomitant inhibition of BKCa channels, may be insufficient to account fully for the [Ca2+ ]i -response to hypoxia.

ILC3 GM-CSF production and mobilisation orchestrate acute intestinal inflammation.

Innate lymphoid cells (ILCs) contribute to host defence and tissue repair but can induce immunopathology. Recent work has revealed tissue-specific roles for ILCs; however, the question of how a small population has large effects on immune homeostasis remains unclear. We identify two mechanisms that ILC3s utilise to exert their effects within intestinal tissue. ILC-driven colitis depends on production of granulocyte macrophage-colony stimulating factor (GM-CSF), which recruits and maintains intestinal inflammatory monocytes. ILCs present in the intestine also enter and exit cryptopatches in a highly dynamic process. During colitis, ILC3s mobilize from cryptopatches, a process that can be inhibited by blocking GM-CSF, and mobilization precedes inflammatory foci elsewhere in the tissue. Together these data identify the IL-23R/GM-CSF axis within ILC3 as a key control point in the accumulation of innate effector cells in the intestine and in the spatio-temporal dynamics of ILCs in the intestinal inflammatory response.

A method for continuous and stable perfusion of tissue and single cell preparations with accurate concentrations of volatile anaesthetics.

BACKGROUND: It is difficult to design a system to reliably deliver volatile anaesthetics such as halothane or isoflurane to in vitro preparations such as tissues or cells cultures: the very volatility of the drugs means that they can rapidly dissipate from even carefully-prepared solutions. Furthermore, many experiments require the control of other gases (such as oxygen or carbon dioxide) which requires constant perfusion. NEW METHOD: We describe a constant perfusion system that is air-tight (i.e., allows the accurate administration of hypoxic or hypercapnic gas mixtures), in which volatile anaesthetic is delivered via calibrated vaporisers by constant bubbling into the perfusing solution (and continuously monitored for stability by infrared spectroscopy in the headspace above the solution). RESULTS: We have confirmed the accuracy (i.e., linear relationship of dissolved concentrations with vapour dial settings) and stability (i.e., over time) of the anaesthetic concentrations in solutions in samples taken from the bottles into which anaesthetic is bubbled, and from samples taken from the tissue perfusion bath, using gas chromatrography-mass spectrometry (GC-MS). CONCLUSIONS: It is possible to deliver volatile anaesthetics in accurate concentrations to cell/tissue preparations whilst adjusting ambient air composition rapidly, stable over sustained time periods.