Determinants of Functional Coupling between Astrocytes and Respiratory Neurons in the Pre-Bötzinger Complex

Respiratory neuronal network activity is thought to require efficient functioning of astrocytes. Here, we analyzed neuron-astrocyte communication in the pre-Bötzinger Complex (preBötC) of rhythmic slice preparations from neonatal mice. In astrocytes that exhibited rhythmic potassium fluxes and glutamate transporter currents, we did not find a translation of respiratory neuronal activity into phase-locked astroglial calcium signals. In up to 20% of astrocytes, 2-photon calcium imaging revealed spontaneous calcium fluctuations, although with no correlation to neuronal activity. Calcium signals could be elicited in preBötC astrocytes by metabotropic glutamate receptor activation or after inhibition of glial glutamate uptake. In the latter case, astrocyte calcium elevation preceded a surge of respiratory neuron discharge activity followed by network failure. We conclude that astrocytes do not exhibit respiratory-rhythmic calcium fluctuations when they are able to prevent synaptic glutamate accumulation. Calcium signaling is, however, observed when glutamate transport processes in astrocytes are suppressed or neuronal discharge activity is excessive

Functional Refinement in the Projection from Ventral Cochlear Nucleus to Lateral Superior Olive Precedes Hearing Onset in Rat

Principal neurons of the lateral superior olive (LSO) compute the interaural intensity differences necessary for localizing high-frequency sounds. To perform this computation, the LSO requires precisely tuned, converging excitatory and inhibitory inputs that are driven by the two ears and that are matched for stimulus frequency. In rodents, the inhibitory inputs, which arise from the medial nucleus of the trapezoid body (MNTB), undergo extensive functional refinement during the first postnatal week. Similar functional refinement of the ascending excitatory pathway, which arises in the anteroventral cochlear nucleus (AVCN), has been assumed but has not been well studied. Using whole-cell voltage clamp in acute brainstem slices of neonatal rats, we examined developmental changes in input strength and pre- and post-synaptic properties of the VCN-LSO pathway. A key question was whether functional refinement in one of the two major input pathways might precede and then guide refinement in the opposite pathway. We find that elimination and strengthening of VCN inputs to the LSO occurs over a similar period to that seen for the ascending inhibitory (MNTB-LSO) pathway. During this period, the fractional contribution provided by NMDA receptors (NMDARs) declines while the contribution from AMPA receptors (AMPARs) increases. In the NMDAR-mediated response, GluN2B-containing NMDARs predominate in the first postnatal week and decline sharply thereafter. Finally, the progressive decrease in paired-pulse depression between birth and hearing onset allows these synapses to follow progressively higher frequencies. Our data are consistent with a model in which the excitatory and inhibitory projections to LSO are functionally refined in parallel during the first postnatal week, and they further suggest that GluN2B-containing NMDARs may mediate early refinement in the VCN-LSO pathway

ICAR: endoscopic skull‐base surgery

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Layer-dependent role of collagen recruitment during loading of the rat bladder wall

In this work, we re-evaluated long-standing conjectures as to the source of the exceptionally large compliance of the bladder wall. Whereas these conjectures were based on indirect measures of loading mechanisms, in this work we take advantage of advances in bioimaging to directly assess collagen fibers and wall architecture during biaxial loading. A custom biaxial mechanical testing system compatible with multiphoton microscopy was used to directly measure the layer-dependent collagen fiber recruitment in bladder tissue from 9 male Fischer rats (4 adult and 5 aged). As for other soft tissues, the bladder loading curve was exponential in shape and could be divided into toe, transition and high stress regimes. The relationship between collagen recruitment and loading curves was evaluated in the context of the inner (lamina propria) and outer (detrusor smooth muscle) layers. The large extensibility of the bladder was found to be possible due to folds in the wall (rugae) that provide a mechanism for low resistance flattening without any discernible recruitment of collagen fibers throughout the toe regime. For more extensible bladders, as the loading extended into the transition regime, a gradual coordinated recruitment of collagen fibers between the lamina propria layer and detrusor smooth muscle layer was found. A second important finding was that wall extensibility could be lost by premature recruitment of collagen in the outer wall that cut short the toe region. This change was correlated with age. This work provides, for the first time, a mechanistic understanding of the role of collagen recruitment in determining bladder extensibility and capacitance

Serotonergic paraneurones in the female mouse urethral epithelium and their potential role in peripheral sensory information processing

The mechanisms underlying detection and transmission of sensory signals arising from visceral organs, such as the urethra, are poorly understood. Recently, specialized ACh-expressing cells embedded in the urethral epithelium have been proposed as chemosensory sentinels for detection of bacterial infection. Here, we examined the morphology and potential role in sensory signalling of a different class of specialized cells that express serotonin (5-HT), termed paraneurones. Urethrae, dorsal root ganglia neurones and spinal cords were isolated from adult female mice and used for immunohistochemistry and calcium imaging. Visceromotor reflexes (VMRs) were recorded in vivo. We identified two morphologically distinct groups of 5-HT+ cells with distinct regional locations: bipolar-like cells predominant in the mid-urethra and multipolar-like cells predominant in the proximal and distal urethra. Sensory nerve fibres positive for calcitonin gene-related peptide, substance P, and TRPV1 were found in close proximity to 5-HT+ paraneurones. In vitro 5-HT (1 μm) stimulation of urethral primary afferent neurones, mimicking 5-HT release from paraneurones, elicited changes in the intracellular calcium concentration ([Ca2+ ]i ) mediated by 5-HT2 and 5-HT3 receptors. Approximately 50% of 5-HT responding cells also responded to capsaicin with changes in the [Ca2+ ]i . In vivo intra-urethral 5-HT application increased VMRs induced by urethral distention and activated pERK in lumbosacral spinal cord neurones. These morphological and functional findings provide insights into a putative paraneurone-neural network within the urethra that utilizes 5-HT signalling, presumably from paraneurones, to modulate primary sensory pathways carrying nociceptive and non-nociceptive (mechano-sensitive) information to the central nervous system