No abstract

The meninges contain adaptive immune cells that provide immunosurveillance of the CNS. These cells are thought to derive from the systemic circulation. Through single-cell analyses, confocal imaging, bone marrow chimeras, and parabiosis experiments, we show that meningeal B cells derive locally from the calvaria, which harbors a bone marrow niche for hematopoiesis. B cells reach the meninges from the calvaria through specialized vascular connections. This calvarial–meningeal path of B cell development may provide the CNS with a constant supply of B cells educated by CNS antigens. Conversely, we show that a subset of antigen-experienced B cells that populate the meninges in aging mice are blood-borne. These results identify a private source for meningeal B cells. which may help maintain immune privilege within the CNS.

Background & Aims-The colonic migrating motor complex (CMMC) is a motor pattern that regulates the movement of fecal matter, through a rhythmic sequence of electrical activity and/or contractions, along the large bowel. CMMCs have largely been studied in empty preparations; we investigated whether local reflexes generated by a fecal pellet modify the CMMC to initiate propulsive activity.

The spontaneous colonic migrating motor complex (CMMC) is a cyclical contractile and electrical event that is the primary motor pattern underlying fecal pellet propulsion along the murine colon. We have combined Ca 2+ imaging with immunohistochemistry to determine the role of different classes of myenteric neurons during the CMMC. Between CMMCs, myenteric neurons usually displayed ongoing but uncoordinated activity. Stroking the mucosa at the oral or anal end of the colon resulted in a CMMC (latency: ∼6 to 10 s; duration: ∼28 s) that consisted of prolonged increases in activity in many myenteric neurons that was correlated to Ca 2+ transients in and displacement of the muscle. These neurons were likely excitatory motor neurons. Activity in individual neurons during the CMMC was similar regardless of whether the CMMC occurred spontaneously or was evoked by anal or oral mucosal stimulation. This suggests that convergent interneuronal pathways exist which generate CMMCs. Interestingly, Ca 2+ transients in a subset of NOS +ve neurons were substantially reduced during the CMMC. These neurons are likely to be inhibitory motor neurons that reduce their activity during a complex (disinhibition) to allow full excitation of the muscle. Local stimulation of the mucosa evoked synchronized Ca 2+ transients in Dogiel Type II (mitotracker/calbindin-positive) neurons after a short delay (∼1-2 s), indicating they were the sensory neurons underlying the CMMC. These local responses were observed in hexamethonium, but were blocked by ondansetron (5-HT 3 antagonist), suggesting Dogiel Type II neurons were activated by 5-HT release from enterochromaffin cells in the mucosa. In fact, removal of the mucosa yielded no spontaneous CMMCs, although many neurons (NOS +ve and NOS −ve) exhibited ongoing activity, including Dogiel Type II neurons. These results suggest that spontaneous or evoked 5-HT release from the mucosa is necessary for the activation of Dogiel Type II neurons that generate CMMCs.

Colonic migrating motor complexes (CMMCs) are neurally mediated, cyclical contractile and electrical events, which typically propagate along the colon every 2-3 min in the mouse. We examined the interactions between myenteric neurons, interstitial cells of Cajal in the myenteric region (ICC-MY) and smooth muscle cells during CMMCs using Ca 2+ imaging. CMMCs occurred spontaneously or were evoked by stimulating the mucosa locally, or by brushing it at either end of the colon. Between CMMCs, most ICC-MY were often quiescent; their lack of activity was correlated with ongoing Ca 2+ transients in varicosities on the axons of presumably inhibitory motor neurons that were on or surrounded ICC-MY. Ca 2+ transients in other varicosities initiated intracellular Ca 2+ waves in adjacent ICC-MY, which were blocked by atropine, suggesting they were on the axons of excitatory motor neurons. Following TTX (1 μm), or blockade of inhibitory neurotransmission with N ω -nitro-l-arginine (l-NA, a NO synthesis inhibitor, 10 μm) and MRS 2500 (a P2Y 1 antagonist, 1 μm), ongoing spark/puff like activity and rhythmic intracellular Ca 2+ waves (38.1 ± 2.9 cycles min −1 ) were observed, yet this activity was uncoupled, even between ICC-MY in close apposition. During spontaneous or evoked CMMCs there was an increase in the frequency (62.9 ± 1.4 cycles min −1 ) and amplitude of Ca 2+ transients in ICC-MY and muscle, which often had synchronized activity. At the same time, activity in varicosites along excitatory and inhibitory motor nerve fibres increased and decreased respectively, leading to an overall excitation of ICC-MY. Atropine (1 μm) reduced the evoked responses in ICC-MY, and subsequent addition of an NK1 antagonist (RP 67580, 500 nm) completely blocked the responses to stimulation, as did applying these drugs in reverse order. An NKII antagonist (MEN 10,376, 500 nm) had no effect on the evoked responses in ICC-MY. Following TTX application, carbachol (1 μm), substance P (1 μm) and an NKI agonist (GR73632, 100 nm) produced the fast oscillations superimposed on a slow increase in Ca 2+ in ICC-MY, whereas SNP (an NO donor, 10 μm) abolished all activity in ICC-MY. In conclusion, ICC-MY, which are under tonic inhibition, are pacemakers whose activity can be synchronized by excitatory nerves to couple the longitudinal and circular muscles during the CMMC. ICC-MY receive excitatory input from motor neurons that release acetylcholine and tachykinins acting on muscarinic and NK1 receptors, respectively.