Biology subjects

Stobart, J. L.

Publications and source records attributed to Stobart, J. L..

4 recordsLinked to original sources

bioRxiv2024

L-Type calcium channels and TRPC3 channels shape brain pericyte calcium signaling and hemodynamics throughout the arteriole to capillary network in vivo

Pericytes play a crucial role in regulating cerebral blood flow (CBF) through processes like vasomotion and neurovascular coupling (NVC). Recent work has identified different pericyte types at distinct points in the cerebrovascular network, such as the arteriole-capillary transition zone (ACT) and distal capillaries, sparking debate about their functional roles in blood flow control. Part of this discussion has comprised the possible mechanisms that may regulate pericyte Ca2+ signaling. Using in vivo two-photon Ca2+ imaging and a pharmacological approach with Ca2+ channel blockers (nimodipine and Pyr3), we assessed the contribution of L-type voltage-gated Ca2+ channels (VGCC) and transient receptor potential canonical 3 (TRPC3) channels to Ca2+ signaling in different pericyte types, ensheathing and capillary pericytes. We also measured local hemodynamics such as vessel diameter, blood cell velocity and flux during vasomotion, and following somatosensory stimulation to evoke NVC. We report that VGCC and TRPC3 channels underlie spontaneous fluctuations in ensheathing pericyte Ca2+ that trigger vasomotor contractions, but the contribution of each of these mechanisms to vascular tone depends on the specific branch of the ACT. Distal capillary pericytes also express L-type VGCCs and TRPC3 channels and they mediate spontaneous Ca2+ signaling in these cells. However, only TRPC3 channels maintain resting capillary tone, possibly by a receptor-operated Ca2+ entry mechanism. By applying the Ca2+ channel blockers during NVC, we found a significant involvement of L-type VGCCs in both pericyte types, influencing their ability to dilate during functional hyperemia. These findings provide new evidence of VGCC and TRPC3 activity in pericytes in vivo and establish a clear distinction between brain pericyte types and their functional roles, opening avenues for innovative strategies to selectively target their Ca2+ dynamics for CBF control. Significance StatementAlthough brain pericytes contribute to the regulation of CBF, there is uncertainty about how different types of pericytes are involved in this process. Ca2+ signaling is believed to be important for the contractility and tone of pericytes, but there is a limited understanding of the Ca2+ pathways in specific pericyte types. Here, we demonstrate that both VGCC and TRPC3 channels are active in distinct types of pericytes throughout the cerebrovascular network, but have different roles in pericyte tone depending on the pericyte location. This has important implications for how pericytes influence vasomotion and neurovascular coupling, which are central processes in CBF regulation. This work also provides the first evidence of TRPC3 channel activity in pericytes in vivo, furthering our understanding of the diverse signaling pathways within these brain mural cells.

neuroscience↗

bioRxiv2023

Cortical astrocyte N-Methyl-D-Aspartate receptors influence whisker barrel activity and sensory discrimination

Cortical astrocytes encode sensory information through their calcium dynamics, but it remains unclear if modulation of astrocyte calcium transients can change somatosensory circuits and behaviour in vivo. Here, we used a novel knockdown approach to selectively reduce astrocyte N-methyl-D-aspartate receptors (NMDAR). We found that these ionotropic receptors contribute to astrocyte Ca2+ transients encoding sensory information. This was essential for the optimal processing of sensory information in nearby neurons, since a reduction in astrocyte NMDARs caused circuit dysfunction and impaired neuronal responses to stimulation. This led to sensory discrimination deficits in the animal. Overall, our findings show that astrocytes can rapidly respond to glutamatergic transmission via their NMDAR and these receptors are an important component for astrocyte-neuron interactions that regulate cortical sensory discrimination in vivo.

neuroscience↗

bioRxiv2020

Differential impact of GABAA Receptors on Inhibitory connectivity for Superficial Pyramidal Neuron Responsiveness In Vivo

A diverse set of GABAA receptors (GABAARs) enable synaptic plasticity adaptations at inhibitory postsynaptic sites in collaboration with the scaffolding protein gephyrin. Early studies helped to identify distinctions between GABAAR subtypes allocated within specific functional circuits, but their contribution to the changing dynamics of a microcircuit remains unclear. Here, using the whisker-barrel system in mouse, we assessed the contribution of specific synaptic GABAAR subtypes and gephyrin scaffolding changes to sensory processing in vivo. We monitored spontaneous and evoked Ca2+ transients in layer 2/3 pyramidal cells with the genetically encoded Ca2+ sensor RCaMP1.07. Using Gabra1 or Gabra2 global and conditional knockout mice, we uncovered that 1- and 2-GABAARs determine the sparseness of L2/3 pyramidal neuron encoding. In a cell-type dependent manner, 1-GABAARs and 2-GABAARs affected neuronal excitability and the reliability of neuronal responses after whisker stimulation. We also discerned that gephyrin with its diverse post-translational modifications (PTMs) shows preference for specific GABAAR subtype to facilitate microcircuit activity. Our results underscore the relevance of the diversity of GABAARs within a cortical microcircuit. Key pointsO_LIWhile GABAergic inhibition from interneuron subtypes regulates cortical microcircuit activity the molecular determinants have remain unclear. C_LIO_LIWe demonstrate that specific-GABAA receptor subtypes contribute differentially to layer 2/3 neuronal activities in mouse barrel cortex. C_LIO_LIImportantly, we link the GABAAR contributions to the scaffolding properties of its important postsynaptic density protein gephyrin. We show that different PTMs on gephyrin determines neuronal excitability via GABAAR recruitment and modulation of inhibition within layer 2/3 neurons. C_LIO_LISpecifically, 1 and 2 subunits containing GABAA receptors, along with their scaffolding protein gephyrin determine the distribution of high, medium and low activity pyramidal neurons during sensory encoding, whereby controlling the total activity of cortical microcircuit. C_LI

neuroscience↗

bioRxiv2020

Distinct signatures of calcium activity in brain pericytes

Even though pericytes have been implicated in various neurological disorders, little is known about their function and signaling pathways in the healthy brain. Here, we characterized cortical pericyte calcium dynamics using two-photon imaging of Pdgfr{beta}-CreERT2;GCaMP6s mice under anesthesia in vivo and in brain slices ex vivo. We found distinct differences between pericyte subtypes in vivo: Ensheathing pericytes exhibited smooth muscle cell-like calcium dynamics, while calcium signals in capillary pericytes were irregular, higher in frequency and occurred in cellular microdomains. In contrast to ensheathing pericytes, capillary pericytes retained their spontaneous calcium signals during prolonged anesthesia and in the absence of blood flow ex vivo. Chemogenetic activation of neurons in vivo and acute increase of extracellular potassium in brain slices strongly decreased calcium activity in capillary pericytes. We propose that neuronal activity-induced elevations in extracellular potassium suppress calcium activity in capillary pericytes, likely mediated by Kir2.2 and KATP channel activation.

neuroscience↗