Homeostatic regulation through strengthening of neuronal network-correlated synaptic inputs.

Homeostatic regulation is essential for stable neuronal function. Several synaptic mechanisms of homeostatic plasticity have been described, but the functional properties of synapses involved in homeostasis are unknown. We used longitudinal two-photon functional imaging of dendritic spine calcium signals in visual and retrosplenial cortices of awake adult mice to quantify the sensory deprivation-induced changes in the responses of functionally identified spines. We found that spines whose activity selectively correlated with intrinsic network activity underwent tumor necrosis factor alpha (TNF-α)-dependent homeostatic increases in their response amplitudes, but spines identified as responsive to sensory stimulation did not. We observed an increase in the global sensory-evoked responses following sensory deprivation, despite the fact that the identified sensory inputs did not strengthen. Instead, global sensory-evoked responses correlated with the strength of network-correlated inputs. Our results suggest that homeostatic regulation of global responses is mediated through changes to intrinsic network-correlated inputs rather than changes to identified sensory inputs thought to drive sensory processing.

The Ups and Downs of Firing Rate Homeostasis.

Torrado Pacheco et al. demonstrate that downward firing rate homeostasis occurs when cellular activity levels increase beyond baseline, but only during sleep-dense periods. In contrast, Hebbian-facilitated changes in firing rate occur independently of sleep and wake states.

Microglia Tweak Retinogeniculate Pathways during Visual Circuit Refinement.

Cheadle et al. reveal that microglia expressing TWEAK facilitate synapse elimination through a novel, non-phagocytic mechanism in the retinogeniculate pathway during visual circuit development. This mechanism is experience-dependent and occurs through the local binding of TWEAK to postsynaptic Fn14.

Deprivation-Induced Homeostatic Spine Scaling In Vivo Is Localized to Dendritic Branches that Have Undergone Recent Spine Loss.

Synaptic scaling is a key homeostatic plasticity mechanism and is thought to be involved in the regulation of cortical activity levels. Here we investigated the spatial scale of homeostatic changes in spine size following sensory deprivation in a subset of inhibitory (layer 2/3 GAD65-positive) and excitatory (layer 5 Thy1-positive) neurons in mouse visual cortex. Using repeated in vivo two-photon imaging, we find that increases in spine size are tumor necrosis factor alpha (TNF-α) dependent and thus are likely associated with synaptic scaling. Rather than occurring at all spines, the observed increases in spine size are spatially localized to a subset of dendritic branches and are correlated with the degree of recent local spine loss within that branch. Using simulations, we show that such a compartmentalized form of synaptic scaling has computational benefits over cell-wide scaling for information processing within the cell.

Interactions between synaptic homeostatic mechanisms: an attempt to reconcile BCM theory, synaptic scaling, and changing excitation/inhibition balance.

Homeostatic plasticity is proposed to be mediated by synaptic changes, such as synaptic scaling and shifts in the excitation/inhibition balance. These mechanisms are thought to be separate from the Bienenstock, Cooper, Munro (BCM) learning rule, where the threshold for the induction of long-term potentiation and long-term depression slides in response to changes in activity levels. Yet, both sets of mechanisms produce a homeostatic response of a relative increase (or decrease) in strength of excitatory synapses in response to overall activity-level changes. Here we review recent studies, with a focus on in vivo experiments, to re-examine the overlap and differences between these two mechanisms and we suggest how they may interact to facilitate firing-rate homeostasis, while maintaining functional properties of neurons.

Integrating Hebbian and homeostatic plasticity: the current state of the field and future research directions.

We summarize here the results presented and subsequent discussion from the meeting on Integrating Hebbian and Homeostatic Plasticity at the Royal Society in April 2016. We first outline the major themes and results presented at the meeting. We next provide a synopsis of the outstanding questions that emerged from the discussion at the end of the meeting and finally suggest potential directions of research that we believe are most promising to develop an understanding of how these two forms of plasticity interact to facilitate functional changes in the brain.This article is part of the themed issue 'Integrating Hebbian and homeostatic plasticity'.

Adult plasticity and cortical reorganization after peripheral lesions.

Following loss of input due to peripheral lesions, functional reorganization occurs in the deprived cortical region in adults. Over a period of hours to months, cells in the lesion projection zone (LPZ) begin to respond to novel stimuli. This reorganization is mediated by two processes: a reduction of inhibition in a gradient throughout the cortex and input remapping via sprouting of axonal arbors from cortical regions spatially adjacent to the LPZ, and strengthening of pre-existing subthreshold inputs. Together these inputs facilitate receptive field remapping of cells in the LPZ. Recent experiments have revealed time courses and potential interactions of the mechanisms associated with functional reorganization, suggesting that large scale reorganization in the adult may utilize plasticity mechanisms prominent during development.

Subnetwork-Specific Homeostatic Plasticity in Mouse Visual Cortex In Vivo.

Homeostatic regulation has been shown to restore cortical activity in vivo following sensory deprivation, but it is unclear whether this recovery is uniform across all cells or specific to a subset of the network. To address this issue, we used chronic calcium imaging in behaving adult mice to examine the activity of individual excitatory and inhibitory neurons in the same region of the layer 2/3 monocular visual cortex following enucleation. We found that only a fraction of excitatory neurons homeostatically recover activity after deprivation and inhibitory neurons show no recovery. Prior to deprivation, excitatory cells that did recover were more likely to have significantly correlated activity with other recovering excitatory neurons, thus forming a subnetwork of recovering neurons. These network level changes are accompanied by a reduction in synaptic inhibition onto all excitatory neurons, suggesting that both synaptic mechanisms and subnetwork activity are important for homeostatic recovery of activity after deprivation.

Nonlinear transfer of signal and noise correlations in cortical networks.

Signal and noise correlations, a prominent feature of cortical activity, reflect the structure and function of networks during sensory processing. However, in addition to reflecting network properties, correlations are also shaped by intrinsic neuronal mechanisms. Here we show that spike threshold transforms correlations by creating nonlinear interactions between signal and noise inputs; even when input noise correlation is constant, spiking noise correlation varies with both the strength and correlation of signal inputs. We characterize these effects systematically in vitro in mice and demonstrate their impact on sensory processing in vivo in gerbils. We also find that the effects of nonlinear correlation transfer on cortical responses are stronger in the synchronized state than in the desynchronized state, and show that they can be reproduced and understood in a model with a simple threshold nonlinearity. Since these effects arise from an intrinsic neuronal property, they are likely to be present across sensory systems and, thus, our results are a critical step toward a general understanding of how correlated spiking relates to the structure and function of cortical networks.

Imaging neuronal populations in behaving rodents: paradigms for studying neural circuits underlying behavior in the mammalian cortex.

Understanding the neural correlates of behavior in the mammalian cortex requires measurements of activity in awake, behaving animals. Rodents have emerged as a powerful model for dissecting the cortical circuits underlying behavior attributable to the convergence of several methods. Genetically encoded calcium indicators combined with viral-mediated or transgenic tools enable chronic monitoring of calcium signals in neuronal populations and subcellular structures of identified cell types. Stable one- and two-photon imaging of neuronal activity in awake, behaving animals is now possible using new behavioral paradigms in head-fixed animals, or using novel miniature head-mounted microscopes in freely moving animals. This mini-symposium will highlight recent applications of these methods for studying sensorimotor integration, decision making, learning, and memory in cortical and subcortical brain areas. We will outline future prospects and challenges for identifying the neural underpinnings of task-dependent behavior using cellular imaging in rodents.

Synaptic scaling and homeostatic plasticity in the mouse visual cortex in vivo.

Homeostatic plasticity is important to maintain a set level of activity in neuronal circuits and has been most extensively studied in cell cultures following activity blockade. It is still unclear, however, whether activity changes associated with mechanisms of homeostatic plasticity occur in vivo, for example after changes in sensory input. Here, we show that activity levels in the visual cortex are significantly decreased after sensory deprivation by retinal lesions, followed by a gradual increase in activity levels in the 48 hr after deprivation. These activity changes are associated with synaptic scaling, manifested in vitro by an increase in mEPSC amplitude and in vivo by an increase in spine size. Together, these data show that homeostatic activity changes occur in vivo in parallel with synaptic scaling.

Three-dimensional imaging of the unsectioned adult spinal cord to assess axon regeneration and glial responses after injury.

Studying regeneration in the central nervous system (CNS) is hampered by current histological and imaging techniques because they provide only partial information about axonal and glial reactions. Here we developed a tetrahydrofuran-based clearing procedure that renders fixed and unsectioned adult CNS tissue transparent and fully penetrable for optical imaging. In large spinal cord segments, we imaged fluorescently labeled cells by 'ultramicroscopy' and two-photon microscopy without the need for histological sectioning. We found that more than a year after injury growth-competent axons regenerated abundantly through the injury site. A few growth-incompetent axons could also regenerate when they bypassed the lesion. Moreover, we accurately determined quantitative changes of glial cells after spinal cord injury. Thus, clearing CNS tissue enables an unambiguous evaluation of axon regeneration and glial reactions. Our clearing procedure also renders other organs transparent, which makes this approach useful for a large number of preclinical paradigms.

Loss of sensory input causes rapid structural changes of inhibitory neurons in adult mouse visual cortex.

A fundamental property of neuronal circuits is the ability to adapt to altered sensory inputs. It is well established that the functional synaptic changes underlying this adaptation are reflected by structural modifications in excitatory neurons. In contrast, the degree to which structural plasticity in inhibitory neurons accompanies functional changes is less clear. Here, we use two-photon imaging to monitor the fine structure of inhibitory neurons in mouse visual cortex after deprivation induced by retinal lesions. We find that a subset of inhibitory neurons carry dendritic spines, which form glutamatergic synapses. Removal of visual input correlates with a rapid and lasting reduction in the number of inhibitory cell spines. Similar to the effects seen for dendritic spines, the number of inhibitory neuron boutons dropped sharply after retinal lesions. Together, these data suggest that structural changes in inhibitory neurons may precede structural changes in excitatory circuitry, which ultimately result in functional adaptation following sensory deprivation.

Molecular and electrophysiological characterization of GFP-expressing CA1 interneurons in GAD65-GFP mice.

The use of transgenic mice in which subtypes of neurons are labeled with a fluorescent protein has greatly facilitated modern neuroscience research. GAD65-GFP mice, which have GABAergic interneurons labeled with GFP, are widely used in many research laboratories, although the properties of the labeled cells have not been studied in detail. Here we investigate these cells in the hippocampal area CA1 and show that they constitute ∼20% of interneurons in this area. The majority of them expresses either reelin (70±2%) or vasoactive intestinal peptide (VIP; 15±2%), while expression of parvalbumin and somatostatin is virtually absent. This strongly suggests they originate from the caudal, and not the medial, ganglionic eminence. GFP-labeled interneurons can be subdivided according to the (partially overlapping) expression of neuropeptide Y (42±3%), cholecystokinin (25±3%), calbindin (20±2%) or calretinin (20±2%). Most of these subtypes (with the exception of calretinin-expressing interneurons) target the dendrites of CA1 pyramidal cells. GFP-labeled interneurons mostly show delayed onset of firing around threshold, and regular firing with moderate frequency adaptation at more depolarized potentials.

Long-term, high-resolution imaging in the mouse neocortex through a chronic cranial window.

To understand the cellular and circuit mechanisms of experience-dependent plasticity, neurons and their synapses need to be studied in the intact brain over extended periods of time. Two-photon excitation laser scanning microscopy (2PLSM), together with expression of fluorescent proteins, enables high-resolution imaging of neuronal structure in vivo. In this protocol we describe a chronic cranial window to obtain optical access to the mouse cerebral cortex for long-term imaging. A small bone flap is replaced with a coverglass, which is permanently sealed in place with dental acrylic, providing a clear imaging window with a large field of view (approximately 0.8-12 mm(2)). The surgical procedure can be completed within approximately 1 h. The preparation allows imaging over time periods of months with arbitrary imaging intervals. The large size of the imaging window facilitates imaging of ongoing structural plasticity of small neuronal structures in mice, with low densities of labeled neurons. The entire dendritic and axonal arbor of individual neurons can be reconstructed.

Glycinergic inhibition in the hippocampus.

Glycine and GABA are the two main inhibitory neurotransmitters in the central nervous system (CNS). While GABA receptors in the hippocampus have been studied in great detail, the role of glycine receptors (GlyRs) in the hippocampus is less understood. Here we examine recent evidence suggesting that GlyRs are present and active throughout the hippocampus. Extracellular glycine levels are controlled through a combination of release and transport mechanisms, both of which, along with the GlyRs themselves, can be modulated by a number of factors. We discuss the role of GlyRs in suppressing excitation by decreasing postsynaptic membrane resistance in the hippocampus, as well as the contribution of GlyRs to both short- and long-term plasticity.

Massive restructuring of neuronal circuits during functional reorganization of adult visual cortex.

The cerebral cortex has the ability to adapt to altered sensory inputs. In the visual cortex, a small lesion to the retina causes the deprived cortical region to become responsive to adjacent parts of the visual field. This extensive topographic remapping is assumed to be mediated by the rewiring of intracortical connections, but the dynamics of this reorganization process remain unknown. We used repeated intrinsic signal and two-photon imaging to monitor functional and structural alterations in adult mouse visual cortex over a period of months following a retinal lesion. The rate at which dendritic spines were lost and gained increased threefold after a small retinal lesion, leading to an almost complete replacement of spines in the deafferented cortex within 2 months. Because this massive remodeling of synaptic structures did not occur when all visual input was removed, it likely reflects the activity-dependent establishment of new cortical circuits that serve the recovery of visual responses.

Cre-dependent expression of multiple transgenes in isolated neurons of the adult forebrain.

BACKGROUND: Transgenic mice with mosaic, Golgi-staining-like expression of enhanced green fluorescent protein (EGFP) have been very useful in studying the dynamics of neuronal structure and function. In order to further investigate the molecular events regulating structural plasticity, it would be useful to express multiple proteins in the same sparse neurons, allowing co-expression of functional proteins or co-labeling of subcellular compartments with other fluorescent proteins. However, it has been difficult to obtain reproducible expression in the same subset of neurons for direct comparison of neurons expressing different functional proteins. PRINCIPAL FINDINGS: Here we describe a Cre-transgenic line that allows reproducible expression of transgenic proteins of choice in a small number of neurons of the adult cortex, hippocampus, striatum, olfactory bulb, subiculum, hypothalamus, superior colliculus and amygdala. We show that using these Cre-transgenic mice, multiple Cre-dependent transgenes can be expressed together in the same isolated neurons. We also describe a Cre-dependent transgenic line expressing a membrane associated EGFP (EGFP-F). Crossed with the Cre-transgenic line, EGFP-F expression starts in the adolescent forebrain, is present in dendrites, dendritic protrusions, axons and boutons and is strong enough for acute or chronic in vivo imaging. SIGNIFICANCE: This triple transgenic approach will aid the morphological and functional characterization of neurons in various Cre-dependent transgenic mice.

Frequency-dependent glycinergic inhibition modulates plasticity in hippocampus.

Previous studies have demonstrated the presence of functional glycine receptors (GlyRs) in hippocampus. In this work, we examine the baseline activity and activity-dependent modulation of GlyRs in region CA1. We find that strychnine-sensitive GlyRs are open in the resting CA1 pyramidal cell, creating a state of tonic inhibition that "shunts" the magnitude of EPSPs evoked by electrical stimulation of the Schaffer collateral inputs. This GlyR-mediated shunting conductance is independent of the presynaptic stimulation rate; however, pairs of presynaptic and postsynaptic action potentials, repeated at frequencies above 5 Hz, reduce the GlyR-mediated conductance and increase peak EPSP magnitudes to levels at least 20% larger than those seen with presynaptic stimulation alone. We refer to this phenomenon as rate-dependent efficacy (RDE). Exogenous GlyR agonists (glycine, taurine) block RDE by preventing the closure of postsynaptic GlyRs. The GlyR antagonist strychnine blocks postsynaptic GlyRs under all conditions, occluding RDE. During RDE, GlyRs are less responsive to local glycine application, suggesting that a reduction in the number or sensitivity of membrane-inserted GlyRs underlies RDE. By extending the RDE induction protocol to include 500 paired presynaptic and postsynaptic spikes, we can induce long-term synaptic depression (LTD). Manipulations that lead to reduced functionality of GlyRs, either pharmacologically or through RDE, also lead to increased LTD. This result suggests that RDE contributes to long-term synaptic plasticity in the hippocampus.