Cortical layers (L) 5 and 6 are populated by intermingled cell-types with distinct inputs and downstream targets. Here, we made optogenetically guided recordings from L5 corticofugal (CF) and L6 ...corticothalamic (CT) neurons in the auditory cortex of awake mice to discern differences in sensory processing and underlying patterns of functional connectivity. Whereas L5 CF neurons showed broad stimulus selectivity with sluggish response latencies and extended temporal non-linearities, L6 CTs exhibited sparse selectivity and rapid temporal processing. L5 CF spikes lagged behind neighboring units and imposed weak feedforward excitation within the local column. By contrast, L6 CT spikes drove robust and sustained activity, particularly in local fast-spiking interneurons. Our findings underscore a duality among sub-cortical projection neurons, where L5 CF units are canonical broadcast neurons that integrate sensory inputs for transmission to distributed downstream targets, while L6 CT neurons are positioned to regulate thalamocortical response gain and selectivity.
Layer 5 (L5) cortical projection neurons innervate far-ranging brain areas to coordinate integrative sensory processing and adaptive behaviors. Here, we characterize a plasticity in L5 auditory ...cortex (ACtx) neurons that innervate the inferior colliculus (IC), thalamus, lateral amygdala and striatum. We track daily changes in sound processing using chronic widefield calcium imaging of L5 axon terminals on the dorsal cap of the IC in awake, adult mice. Sound level growth functions at the level of the auditory nerve and corticocollicular axon terminals are both strongly depressed hours after noise-induced damage of cochlear afferent synapses. Corticocollicular response gain rebounded above baseline levels by the following day and remained elevated for several weeks despite a persistent reduction in auditory nerve input. Sustained potentiation of excitatory ACtx projection neurons that innervate multiple limbic and subcortical auditory centers may underlie hyperexcitability and aberrant functional coupling of distributed brain networks in tinnitus and hyperacusis.
The calibration of complex computer codes using uncertainty quantification (UQ) methods is a rich area of statistical methodological development. When applying these techniques to simulators with ...spatial output, it is now standard to use principal component decomposition to reduce the dimensions of the outputs in order to allow Gaussian process emulators to predict the output for calibration. We introduce the "terminal case," in which the model cannot reproduce observations to within model discrepancy, and for which standard calibration methods in UQ fail to give sensible results. We show that even when there is no such issue with the model, the standard decomposition on the outputs can and usually does lead to a terminal case analysis. We present a simple test to allow a practitioner to establish whether their experiment will result in a terminal case analysis, and a methodology for defining calibration-optimal bases that avoid this whenever it is not inevitable. We present the optimal rotation algorithm for doing this, and demonstrate its efficacy for an idealized example for which the usual principal component methods fail. We apply these ideas to the CanAM4 model to demonstrate the terminal case issue arising for climate models. We discuss climate model tuning and the estimation of model discrepancy within this context, and show how the optimal rotation algorithm can be used in developing practical climate model tuning tools.
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Abstract
The mouse auditory cortex (ACtx) contains two core fields—primary auditory cortex (A1) and anterior auditory field (AAF)—arranged in a mirror reversal tonotopic gradient. The best frequency ...(BF) organization and naming scheme for additional higher order fields remain a matter of debate, as does the correspondence between smoothly varying global tonotopy and heterogeneity in local cellular tuning. Here, we performed chronic widefield and two-photon calcium imaging from the ACtx of awake Thy1-GCaMP6s reporter mice. Data-driven parcellation of widefield maps identified five fields, including a previously unidentified area at the ventral posterior extreme of the ACtx (VPAF) and a tonotopically organized suprarhinal auditory field (SRAF) that extended laterally as far as ectorhinal cortex. Widefield maps were stable over time, where single pixel BFs fluctuated by less than 0.5 octaves throughout a 1-month imaging period. After accounting for neuropil signal and frequency tuning strength, BF organization in neighboring layer 2/3 neurons was intermediate to the heterogeneous salt and pepper organization and the highly precise local organization that have each been described in prior studies. Multiscale imaging data suggest there is no ultrasonic field or secondary auditory cortex in the mouse. Instead, VPAF and a dorsal posterior (DP) field emerged as the strongest candidates for higher order auditory areas.
Corticothalamic (CT) neurons comprise the largest component of the descending sensory corticofugal pathway, but their contributions to brain function and behavior remain an unsolved mystery. To ...address the hypothesis that layer 6 (L6) CTs may be activated by extra-sensory inputs prior to anticipated sounds, we performed optogenetically targeted single-unit recordings and two-photon imaging of Ntsr1-Cre+ L6 CT neurons in the primary auditory cortex (A1) while mice were engaged in an active listening task. We found that L6 CTs and other L6 units began spiking hundreds of milliseconds prior to orofacial movements linked to sound presentation and reward, but not to other movements such as locomotion, which were not linked to an explicit behavioral task. Rabies tracing of monosynaptic inputs to A1 L6 CT neurons revealed a narrow strip of cholinergic and non-cholinergic projection neurons in the external globus pallidus, suggesting a potential source of motor-related input. These findings identify new pathways and local circuits for motor modulation of sound processing and suggest a new role for CT neurons in active sensing.
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•L6 corticothalamic neurons (L6 CTs) were isolated during active listening tasks•L6 CT activity increases prior to movements that trigger sound and reward•Motor corollary input activates L6 CTs at similar latency to FS interneurons•Rabies tracing reveals monosynaptic inputs onto L6 CTs from globus pallidus
Corticothalamic neurons (CTs) regulate excitability throughout the thalamocortical loop, but what regulates their excitability? Clayton et al. record from mouse auditory layer 6 CTs during active listening tasks. They find strong motor-related extra-sensory activation of layer 6 CTs beginning hundreds of milliseconds prior to movement onset.
Active search is a ubiquitous goal-driven behavior wherein organisms purposefully investigate the sensory environment to locate a target object. During active search, brain circuits analyze a stream ...of sensory information from the external environment, adjusting for internal signals related to self-generated movement or “top-down” weighting of anticipated target and distractor properties. Sensory responses in the cortex can be modulated by internal state 1–9, though the extent and form of modulation arising in the cortex de novo versus an inheritance from subcortical stations is not clear 4, 8–12. We addressed this question by simultaneously recording from auditory and visual regions of the thalamus (MG and LG, respectively) while mice used dynamic auditory or visual feedback to search for a hidden target within an annular track. Locomotion was associated with strongly suppressed responses and reduced decoding accuracy in MG but a subtle increase in LG spiking. Because stimuli in one modality provided critical information about target location while the other served as a distractor, we could also estimate the importance of task relevance in both thalamic subdivisions. In contrast to the effects of locomotion, we found that LG responses were reduced overall yet decoded stimuli more accurately when vision was behaviorally relevant, whereas task relevance had little effect on MG responses. This double dissociation between the influences of task relevance and movement in MG and LG highlights a role for extrasensory modulation in the thalamus but also suggests key differences in the organization of modulatory circuitry between the auditory and visual pathways.
•Auditory thalamic activity is significantly suppressed during movement•Visual thalamic activity is subtly enhanced, only at high running speeds•Behavioral relevance modulates activity in visual—but not auditory—thalamus•Locomotor state and task relevance can be used to improve neural decoding accuracy
Williamson et al. recorded from the auditory and visual thalamus of mice engaged in an audiovisual search task. They find a double dissociation between task relevance and movement, highlighting a role for modulation of thalamic responses by internal state and suggesting key differences in modulatory circuitry between auditory and visual pathways.
In sensory systems, representational features of increasing complexity emerge at successive stages of processing. In the mammalian auditory pathway, the clearest change from brainstem to cortex is ...defined by what is lost, not by what is gained, in that high-fidelity temporal coding becomes increasingly restricted to slower acoustic modulation rates.1,2 Here, we explore the idea that sluggish temporal processing is more than just an inability for fast processing, but instead reflects an emergent specialization for encoding sound features that unfold on very slow timescales.3,4 We performed simultaneous single unit ensemble recordings from three hierarchical stages of auditory processing in awake mice – the inferior colliculus (IC), medial geniculate body of the thalamus (MGB) and primary auditory cortex (A1). As expected, temporal coding of brief local intervals (0.001 – 0.1 s) separating consecutive noise bursts was robust in the IC and declined across MGB and A1. By contrast, slowly developing (∼1 s period) global rhythmic patterns of inter-burst interval sequences strongly modulated A1 spiking, were weakly captured by MGB neurons, and not at all by IC neurons. Shifts in stimulus regularity were not represented by changes in A1 spike rates, but rather in how the spikes were arranged in time. These findings show that low-level auditory neurons with fast timescales encode isolated sound features but not the longer gestalt, while the extended timescales in higher-level areas can facilitate sensitivity to slower contextual changes in the sensory environment.
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•Simultaneous unit recordings from the mouse auditory midbrain, thalamus, and cortex•Subcortical stations decode short temporal intervals, not longer rhythmic patterns•Cortical units decode slower changes in stimulus patterns, not short intervals•Changes in stimulus regularity are reflected in cortical spike timing, not rate
Asokan et al. show an emergent sensitivity for slow temporal rhythms at the level of the auditory cortex. Whereas single units in the mouse midbrain and thalamus encode local temporal intervals with high fidelity, cortical units integrate over extended periods to distinguish between random or regular interval sequences by adjusting spike timing.
The Notch-signaling pathway is normally activated by Notch–ligand interactions. A recent structural analysis suggested that a novel O-linked hexose modification on serine 435 of the mammalian NOTCH1 ...core ligand-binding domain lies at the interface with its ligands. This serine occurs between conserved cysteines 3 and 4 of Epidermal Growth Factor-like (EGF) repeat 11 of NOTCH1, a site distinct from those modified by protein O-glucosyltransferase 1 (POGLUT1), suggesting that a different enzyme is responsible. Here, we identify two novel protein O-glucosyltransferases, POGLUT2 and POGLUT3 (formerly KDELC1 and KDELC2, respectively), which transfer O-glucose (O-Glc) from UDP-Glc to serine 435. Mass spectrometric analysis of NOTCH1 produced in HEK293T cells lacking POGLUT2, POGLUT3, or both genes showed that either POGLUT2 or POGLUT3 can add this novel O-Glc modification. EGF11 of NOTCH2 does not have a serine residue in the same location for this O-glucosylation, but EGF10 of NOTCH3 (homologous to EGF11 in NOTCH1 and -2) is also modified at the same position. Comparison of the sites suggests a consensus sequence for modification. In vitro assays with POGLUT2 and POGLUT3 showed that both enzymes modified only properly folded EGF repeats and displayed distinct acceptor specificities toward NOTCH1 EGF11 and NOTCH3 EGF10. Mutation of the O-Glc modification site on EGF11 (serine 435) in combination with sensitizing O-fucose mutations in EGF8 or EGF12 affected cell-surface presentation of NOTCH1 or reduced activation of NOTCH1 by Delta-like1, respectively. This study identifies a previously undescribed mechanism for fine-tuning the Notch-signaling pathway in mammals.
O-glycosylation of Epidermal Growth Factor-like (EGF) repeats plays crucial roles in protein folding, trafficking and function. The Notch extracellular domain has been used as a model to study these ...mechanisms due to its many O-glycosylated EGF repeats. Three enzymes were previously known to O-glycosylate Notch EGF repeats: Protein O-Glucosyltransferase 1 (POGLUT1), Protein O-Fucosyltransferase 1 (POFUT1), and EGF Domain Specific O-Linked N-Acetylglucosamine Transferase (EOGT). All of these modifications affect Notch activity. Recently, POGLUT2 and POGLUT3 were identified as two novel O-glucosyltransferases that modify a few Notch EGF repeats at sites distinct from those modified by POGLUT1. Comparison of these modification sites revealed a putative consensus sequence which predicted modification of many extracellular matrix proteins including fibrillins (FBNs) and Latent TGFβ-binding proteins (LTBPs). Glycoproteomic analysis revealed that approximately half of the 47 EGF repeats in FBN1 and FBN2, and half of the 18 EGF repeats in LTBP1, are modified by POGLUT2 and/or POGLUT3. Cellular assays showed that loss of modifications by POGLUT2 and/or POGLUT3 significantly reduces FBN1 secretion. There is precedent for EGF modifications to affect protein-protein interactions, as has been demonstrated by research of POGLUT1 and POFUT1 modifications on Notch. Here we discuss the identification and characterization of POGLUT2 and POGLUT3 and the ongoing research that continues to elucidate the biological significance of these novel enzymes.
Fibrillin-1 (FBN1) is the major component of extracellular matrix microfibrils, which are required for proper development of elastic tissues, including the heart and lungs. Through protein–protein ...interactions with latent transforming growth factor (TGF) β-binding protein 1 (LTBP1), microfibrils regulate TGF-β signaling. Mutations within the 47 epidermal growth factor-like (EGF) repeats of FBN1 cause autosomal dominant disorders including Marfan Syndrome, which is characterized by disrupted TGF-β signaling. We recently identified two novel protein O-glucosyltransferases, Protein O-glucosyltransferase 2 (POGLUT2) and 3 (POGLUT3), that modify a small fraction of EGF repeats on Notch. Here, using mass spectral analysis, we show that POGLUT2 and POGLUT3 also modify over half of the EGF repeats on FBN1, fibrillin-2 (FBN2), and LTBP1. While most sites are modified by both enzymes, some sites show a preference for either POGLUT2 or POGLUT3. POGLUT2 and POGLUT3 are homologs of POGLUT1, which stabilizes Notch proteins by addition of O-glucose to Notch EGF repeats. Like POGLUT1, POGLUT2 and 3 can discern a folded versus unfolded EGF repeat, suggesting POGLUT2 and 3 are involved in a protein folding pathway. In vitro secretion assays using the N-terminal portion of recombinant FBN1 revealed reduced FBN1 secretion in POGLUT2 knockout, POGLUT3 knockout, and POGLUT2 and 3 double-knockout HEK293T cells compared with wild type. These results illustrate that POGLUT2 and 3 function together to O-glucosylate protein substrates and that these modifications play a role in the secretion of substrate proteins. It will be interesting to see how disease variants in these proteins affect their O-glucosylation.
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