Biology subjects

Norton, A.

Publications and source records attributed to Norton, A..

5 recordsLinked to original sources

bioRxiv2025

Purkinje cell collaterals preferentially target a subtype of molecular layer interneuron

In addition to providing outputs from the cerebellar cortex, Purkinje cell (PC) axon collaterals target other PCs, molecular layer interneurons (MLIs), and Purkinje layer interneurons (PLIs). It was recently shown that MLIs consist of two subtypes, but the properties of PC synapses onto these subtypes was not known and it was assumed that all PC collateral to MLI synapses would provide positive feedback to PCs. Clarifying the PC connectivity onto MLI subtypes is vital to understating the influence of feedback from PC collaterals because MLI1s primarily inhibit PCs whereas MLI2s mainly inhibit MLI1s and disinhibit PCs. Here we use a combination of serial EM and optogenetic studies to characterize PC synapses onto MLI subtypes in mice. EM reconstructions show that PCs make 53% of their synapses onto other PCs, 32% onto PLIs, 6% onto MLI1s and 7% onto MLI2s. Since there are far more MLI1s than MLI2s, each MLI2 is expected to receive many more synapses than each MLI1. In slice experiments, optogenetic activation of PCs evokes inhibitory currents in most MLI2s, but primarily disinhibits MLI1s. We also find that candelabrum cells, a type of PLI, form many more synapses onto MLI1s than MLI2s. It is therefore expected that both PC-MLI2-MLI1-PC and PC-PLI-MLI1-PC pathways allow increased PC firing to disinhibit MLI1s, which are known to reduce dendritic PC calcium signals and suppress plasticity at granule cell to PC synapses. These pathways provide negative feedback that act in concert with PC-PC synapses to counter elevations in PC firing. Significance StatementPurkinje cells (PCs) influence processing by inhibiting neurons in the cerebellar cortex, including other PCs, molecular layer interneurons (MLIs) and Purkinje layer interneurons (PLIs). The influence of PC-MLI synapses is not known because there are recently identified MLI subtypes with opposing effects: MLI1s inhibit PCs whereas MLI2s inhibit MLI1s and disinhibit PCs. We used serial EM and optogenetic studies to characterize PC synapses onto MLI subtypes and found that PCs preferentially inhibit MLI2s and disinhibit MLI1s. We also found that candelabrum cells (a type of PLI) preferentially inhibit MLI1s. These findings establish that PC-PC synapses, the PC-MLI2-MLI1-PC pathway and the PC-candelabrum cell-MLI1-PC pathway act together to allow alterations in PC firing to provide negative feedback to other PCs.

neuroscience↗

bioRxiv2025

Climbing fibers selectively recruit disinhibitory interneurons to enhance dendritic calcium signaling in cerebellar Purkinje cells

Climbing fiber (CF) inputs to Purkinje cells (PCs) instruct plasticity and learning in the cerebellum1-3. Paradoxically, CFs also excite molecular layer interneurons (MLIs)4,5, a cell-type that inhibits PCs and can restrict plasticity and learning6,7. However, two types of MLIs with opposing influences have recently been identified: MLI1s inhibit PCs, reduce dendritic calcium signals, and suppress plasticity of granule cell to PC synapses2,6-9, whereas MLI2s inhibit MLI1s and disinhibit PCs8. To determine how CFs can activate MLIs without also suppressing the PC calcium signals necessary for plasticity and learning, we investigated the specificity of CF inputs onto MLIs. Serial EM reconstructions indicate that CFs contact both MLI subtypes without making conventional synapses, but more CFs contact each MLI2 via more sites with larger contact areas. Slice experiments indicate that CFs preferentially excite MLI2s via glutamate spillover4,5. In agreement with these anatomical and slice experiments, in vivo Neuropixels recordings show that spontaneous CF activity excites MLI2s, inhibits MLI1s, and disinhibits PCs. In contrast, learning-related sensory stimulation produced more complex responses, driving convergent CF and granule cell inputs that could either activate or suppress MLI1s. This balance was robustly shifted toward MLI1 suppression when CFs were synchronously active, in turn elevating the PC dendritic calcium signals necessary for LTD. These data provide mechanistic insight into why CF synchrony can be highly effective at inducing cerebellar learning2,3 by revealing a critical disinhibitory circuit that allows CFs to act through MLIs to enhance PC dendritic calcium signals necessary for plasticity.

neuroscience↗

bioRxiv2025

Mesenteric ischemia and bacterial translocation precipitate the intoxication phase of yellow fever

Yellow fever (YF) is classically conceptualized as a hepatotropic disease; indeed, the liver is the primary site of yellow fever virus (YFV) replication. However, circumstantial evidence suggests that extra-hepatic disease may be important for the [~]30% of YF cases that progress to the severe "intoxication" phase of the disease. Using a Syrian hamster-adapted (HA)-YFV, we worked backwards from observations in humans to examine early events that precipitate the intoxication phase of YF. HA-YFV caused severe disease in [~]80% of infected animals characterized by lethargy and weight loss that progressed to widespread petechiae and death by day 6. Clinical chemistry, coagulation testing, histology, immunohistochemistry, and in-situ hybridization were consistent with a cascade of hepatocyte-specific virus replication causing liver damage and a defect in clotting factor synthesis. Despite a lack of extra-hepatic HA-YFV replication, severe pathology was observed in the intestines and pancreas. Histopathological analysis over the time-course of HA-YFV infection revealed an ischemic pattern in these tissues, culminating in fibrinoid/coagulative necrosis of these organs. Further investigation showed that ischemia-induced erosion of the gut epithelial barrier serves as an entry point for luminal bacteria that spread systemically via the portal system. Thus, the intoxication phase of YF is a sepsis-like syndrome caused by translocation of bacteria from a damaged gastrointestinal tract. Evaluation of human YF cases for these previously overlooked disease features confirmed this overarching mechanism: bacteria were identified in the portal vein and liver parenchyma of fatal YF cases along with elevations in plasma markers of bacteremia and a bacteria-driven inflammatory response. Importantly, blood concentrations of the gastrointestinal damage marker intestinal fatty acid binding protein (I-FABP) were significantly elevated in fatal YF cases relative to non-fatal cases, suggesting that I-FABP measurements could be useful in prognosis and treatment decision making. Our findings tie together several recent and historically unexplained observations surrounding the highly-lethal intoxication phase of YF in humans: a high AST/ALT ratio, "black vomit," pancreatitis, and paradoxical neutrophilia. A better appreciation for the drivers of mesenteric ischemia, and preemption of bacterial sepsis, may improve outcomes in cases of severe YF.

microbiology↗

bioRxiv2024

Processing reliant on granule cells is essential for motor learning but dispensable for many cerebellar-dependent behaviors

Cerebellar dysfunction leads to motor, learning, emotional, and social deficits. It is assumed that these deficits arise from impaired processing of mossy fiber inputs that activate granule cells (GCs) that in turn excite Purkinje cells (PCs). However, high-frequency spontaneous PC firing might also influence behaviors. To clarify how the cerebellum regulates behaviors, we compared the effects of disrupting either GC signaling, which selectively perturbs cerebellar processing, or PC signaling, which disrupts cerebellar processing and spontaneous PC firing. We find that both GC and PC signaling are required for eyeblink conditioning and vestibulo-ocular reflex (VOR) learning. However, disrupting PC signaling impairs baseline VOR, anxiety, and social behaviors, but abolishing GC signaling does not. This establishes that cerebellar processing is essential for motor learning, but is not required for many cerebellum-dependent behaviors. This suggests that such behaviors could be rescued by elevating firing in downstream targets, as shown previously for social deficits.

neuroscience↗

bioRxiv2023

Cerebellar circuits for disinhibition and synchronous inhibition

The cerebellar cortex contributes to diverse behaviors by transforming mossy fiber inputs into predictions in the form of Purkinje cell (PC) outputs, and then refining those predictions1. Molecular layer interneurons (MLIs) account for approximately 80% of the inhibitory interneurons in the cerebellar cortex2, and are vital to cerebellar processing1,3. MLIs are thought to primarily inhibit PCs and suppress the plasticity of excitatory synapses onto PCs. MLIs also inhibit, and are electrically coupled to, other MLIs4-7, but the functional significance of these connections is not known1,3. Behavioral studies suggest that cerebellar-dependent learning is gated by disinhibition of PCs, but the source of such disinhibition has not been identified8. Here we find that two recently recognized MLI subtypes2, MLI1 and MLI2, have highly specialized connectivity that allows them to serve very different functional roles. MLI1s primarily inhibit PCs, are electrically coupled to each other, fire synchronously with other MLI1s on the millisecond time scale in vivo, and synchronously pause PC firing. MLI2s are not electrically coupled, they primarily inhibit MLI1s and disinhibit PCs, and are well suited to gating cerebellar-dependent learning8. These findings require a major reevaluation of processing within the cerebellum in which disinhibition, a powerful circuit motif present in the cerebral cortex and elsewhere9-17, greatly increases the computational power and flexibility of the cerebellum. They also suggest that millisecond time scale synchronous firing of electrically-coupled MLI1s helps regulate the output of the cerebellar cortex by synchronously pausing PC firing, which has been shown to evoke precisely-timed firing in PC targets18.

neuroscience↗