PhDs in Press: Winter/Spring 2014
/The drought that afflicted Calfornia this winter was in no way mirrored in the publications authored by members of the Stanford Neurosciences PhD Program.
This Winter/Spring saw 10 PhD students publishing first-author papers (Lief and Joanna, Egle, Kira, Sergio and Corbett, David K, Poh Hui, Mridu and Nathan). They were joined in authorial success by an additional 13 graduate researchers who were 2nd-nth authors (Aslihan, Logan, Kelly, Ivan, Astra, Greg, Izumi, Georgia, Nick Steinmetz, Jake, Tina, Hannah and Mark; not to mention Kira and Poh Hui who also had 2nd-nth author papers).
Witness the majestic variety of neuroscience research being done by the Stanford Neurosciences Graduate Community. The Diesseroth lab makes genetically encoded tools that use Boolean logic. The science partnership of Corbett Bennett and Sergio Arroyo continues with a review article on nicotinic modulation of cortical circuits. Paul Buckmaster's lab publishes a study of epileptic sea lions off the California coast. And so much more...
Continue below for a full list of the articles (complete with links and abstracts).
Targeting cells with single vectors using multiple-feature Boolean logic
Lief Fenno*, Joanna Mattis*, Charu Ramakrishnan*, Minsuk Hyun, Soo Yeun Lee, Miao He, Jason Tucciarone, Aslihan Selimbeyoglu, Andre Berndt, Logan Grosenick, Kelly Zalocusky, Hannah Bernstein, Haley Swanson, Chelsey Perry, Ilka Diester, Frederick Boyce, Caroline Bass, Rachel Never, Z Josh Huang and Karl Deisseroth.
“Precisely defining the roles of specific cell types is an intriguing frontier in the study of intact biological systems and has stimulated the rapid development of genetically encoded tools for observation and control. However, targeting these tools with adequate specificity remains challenging: most cell types are best defined by the intersection of two or more features such as active promoter elements, location and connectivity. Here we have combined engineered introns with specific recombinases to achieve expression of genetically encoded tools that is conditional upon multiple cell-type features, using Boolean logical operations all governed by a single versatile vector. We used this approach to target intersectionally specified populations of inhibitory interneurons in mammalian hippocampus and neurons of the ventral tegmental area defined by both genetic and wiring properties. This flexible and modular approach may expand the application of genetically encoded interventional and observational tools for intact-systems biology”
Astrocytic TGF-β Signaling Limits Inflammation and Reduces Neuronal Damage during Central Nervous System Toxoplasma Infection
Egle Cekanaviciute, Hans Dietrich, Robert Axtell, Aaron Williams, Riann Egusquiza, Karen Wai, Anita Koshy and Marion Buckwalter.
“The balance between controlling infection and limiting inflammation is particularly precarious in the brain because of its unique vulnerability to the toxic effects of inflammation. Astrocytes have been implicated as key regulators of neuroinflammation in CNS infections, including infection with Toxoplasma gondii, a protozoan parasite that naturally establishes a chronic CNS infection in mice and humans. In CNS toxoplasmosis, astrocytes are critical to controlling parasite growth. They secrete proinflammatory cytokines and physically encircle parasites. However, the molecular mechanisms used by astrocytes to limit neuroinflammation during toxoplasmic encephalitis have not yet been identified. TGF-β signaling in astrocytes is of particular interest because TGF-β is universally upregulated during CNS infection and serves master regulatory and primarily anti-inflammatory functions. We report in this study that TGF-β signaling is activated in astrocytes during toxoplasmic encephalitis and that inhibition of astrocytic TGF-β signaling increases immune cell infiltration, uncouples proinflammatory cytokine and chemokine production from CNS parasite burden, and increases neuronal injury. Remarkably, we show that the effects of inhibiting astrocytic TGF-β signaling are independent of parasite burden and the ability of GFAP+ astrocytes to physically encircle parasites.”
Microglial dysfunction in brain aging and Alzheimer's disease.
Kira Mosher and Tony Wyss-Coray.
“Microglia, the immune cells of the central nervous system, have long been a subject of study in the Alzheimer’s disease (AD) field due to their dramatic responses to the pathophysiology of the disease. With several large-scale genetic studies in the past year implicating microglial molecules in AD, the potential significance of these cells has become more prominent than ever before. As a disease that is tightly linked to aging, it is perhaps not entirely surprising that microglia of the AD brain share some phenotypes with aging microglia. Yet the relative impacts of both conditions on microglia are less frequently considered in concert. Furthermore, microglial “activation” and “neuroinflammation” are commonly analyzed in studies of neurodegeneration but are somewhat ill-defined concepts that in fact encompass multiple cellular processes. In this review, we have enumerated six distinct functions of microglia and discuss the specific effects of both aging and AD. By calling attention to the commonalities of these two states, we hope to inspire new approaches for dissecting microglial mechanisms.”
Nicotinic Modulation of Cortical Circuits.
Sergio Arroyo*, Corbett Bennett* and Shaul Hestrin.
“The ascending cholinergic neuromodulatory system sends projections throughout cortex and has been shown to play an important role in a number of cognitive functions including arousal, working memory, and attention. However, despite a wealth of behavioral and anatomical data, understanding how cholinergic synapses modulate cortical function has been limited by the inability to selectively activate cholinergic axons. Now, with the development of optogenetic tools and cell-type specific Cre-driver mouse lines, it has become possible to stimulate cholinergic axons from the basal forebrain (BF) and probe cholinergic synapses in the cortex for the first time. Here we review recent work studying the cell-type specificity of nicotinic signaling in the cortex, synaptic mechanisms mediating cholinergic transmission, and the potential functional role of nicotinic modulation.”
Insights from the retina into the diverse and general computations of adaptation, detection, and prediction.
David Kastner and Steve Baccus
“The retina performs a diverse set of complex, nonlinear, computations, beyond the simple linear photoreceptor weighting assumed in the classical understanding of ganglion cell receptive fields. Here we attempt to organize these computations and extract rules that correspond to three distinct goals of early sensory systems. First, the retina acts efficiently to transmit information to the higher brain for further processing. We observe that although the retina adapts to a number of complex statistics, many of these may be explained by local adaptation to the mean signal strength at that stage. Second, ganglion cells signal the detection of a diverse set of features. Recent results indicate that feature selectivity arises through the action of specific inhibition, rather than through the convergence of excitation as in classical cortical models. Finally, the retina conveys predictions about the stimulus, a function usually attributed only to the higher brain. We expect that computational and mechanistic rules associated with these classes of functions will be an important guide in the study of other neural circuits.”
Local F-actin network links synapse formation and axon branching.
Poh Hui Chia, Baoyu Chen, Pengpeng Li, Michael Rosen, Kang Shen.
“Axonal branching and synapse formation are tightly linked developmental events during the establishment of synaptic circuits. Newly formed synapses promote branch initiation and stability. However, little is known about molecular mechanisms that link these two processes. Here, we show that local assembly of an F-actin cytoskeleton at nascent presynaptic sites initiates both synapse formation and axon branching. We further find that assembly of the F-actin network requires a direct interaction between the synaptic cell adhesion molecule SYG-1 and a key regulator of actin cytoskeleton, the WVE-1/WAVE regulatory complex (WRC). SYG-1 cytoplasmic tail binds to the WRC using a consensus WRC interacting receptor sequence (WIRS). WRC mutants or mutating the SYG-1 WIRS motif leads to loss of local F-actin, synaptic material, and axonal branches. Together, these data suggest that synaptic adhesion molecules, which serve as a necessary component for both synaptogenesis and axonal branch formation, directly regulate subcellular actin cytoskeletal organization.”
A SxIP motif interaction at the microtubule plus end is important for processive retrograde axonal transport.
Mridu Kapur, Michael Maloney, Wei Wang, Xinyu Chen, Ivan Millan, Trevor Mooney, Jie Yang, Yanmin Yang.
“The retrograde transport of endosomes within axons proceeds with remarkable uniformity despite having to navigate a discontinuous microtubule network. The mechanisms through which this navigation is achieved remain elusive. In this report, we demonstrate that access of SxIP motif proteins, such as BPAG1n4, to the microtubule plus end is important for the maintenance of processive and sustained retrograde transport along the axon. Disruption of this interaction at the microtubule plus end significantly increases endosome stalling. Our study thus provides strong insight into the role of plus-end-binding proteins in the processive navigation of cargo within the axon.”
Spatially reciprocal inhibition of inhibition within a stimulus selection network in the avian midbrain.
C Alex Goddard, Shreesh Mysore, Astra Bryant, John Huguenard, Eric Knudsen
“Reciprocal inhibition between inhibitory projection neurons has been proposed as the most efficient circuit motif to achieve the flexible selection of one stimulus among competing alternatives. However, whether such a motif exists in networks that mediate selection is unclear. Here, we study the connectivity within the nucleus isthmi pars magnocellularis (Imc), a GABAergic nucleus that mediates competitive selection in the midbrain stimulus selection network. Using laser photostimulation of caged glutamate, we find that feedback inhibitory connectivity is global within the Imc. Unlike typical lateral inhibition in other circuits, intra-Imc inhibition remains functionally powerful over long distances. Anatomically, we observed long-range axonal projections and retrograde somatic labeling from focal injections of bi-directional tracers in the Imc, consistent with spatial reciprocity of intra-Imc inhibition. Together, the data indicate that spatially reciprocal inhibition of inhibition occurs throughout the Imc. Thus, the midbrain selection circuit possesses the most efficient circuit motif possible for fast, reliable, and flexible selection.”
Young blood reverses age-related impairments in cognitive function and synaptic plasticity in mice.
Saul Villeda, Kristopher Plambeck, Jinte Middledorp, Joseph Castellano, Kira Mosher, Jian Luo, Lucas Smith, Gregor Bieri, Karin Lin, Daniela Berdnik, Rafael Wabl, Joe Udeochu, Elizabeth Wheatley, Bende Zuo, Danielle Simmons, Xinmin Xie, Frank Longo and Tony Wyss-Coray.
“As human lifespan increases, a greater fraction of the population is suffering from age-related cognitive impairments, making it important to elucidate a means to combat the effects of aging. Here we report that exposure of an aged animal to young blood can counteract and reverse pre-existing effects of brain aging at the molecular, structural, functional and cognitive level. Genome-wide microarray analysis of heterochronic parabionts-in which circulatory systems of young and aged animals are connected-identified synaptic plasticity-related transcriptional changes in the hippocampus of aged mice. Dendritic spine density of mature neurons increased and synaptic plasticity improved in the hippocampus of aged heterochronic parabionts. At the cognitive level, systemic administration of young blood plasma into aged mice improved age-related cognitive impairments in both contextual fear conditioning and spatial learning and memory. Structural and cognitive enhancements elicited by exposure to young blood are mediated, in part, by activation of the cyclic AMP response element binding protein (Creb) in the aged hippocampus. Our data indicate that exposure of aged mice to young blood late in life is capable of rejuvenating synaptic plasticity and improving cognitive function.”
Hippocampal neuropathology of domoic acid-induced epilepsy in California sea lions (Zalophus californianus).
Paul Buckmaster, Xiling Wen, Izumi Toyoda, Frances Gulland, Willian Van Bonn.
“California sea lions (Zalophus californianus) are abundant human-sized carnivores with large gyrencephalic brains. They develop epilepsy after experiencing status epilepticus when naturally exposed to domoic acid. We tested whether sea lions previously exposed to DA (chronic DA sea lions) display hippocampal neuropathology similar to that of human patients with temporal lobe epilepsy. Hippocampi were obtained from control and chronic DA sea lions. Stereology was used to estimate numbers of Nissl-stained neurons per hippocampus in the granule cell layer, hilus, and pyramidal cell layer of CA3, CA2, and CA1 subfields. Adjacent sections were processed for somatostatin immunoreactivity or Timm-stained, and the extent of mossy fiber sprouting was measured stereologically. Chronic DA sea lions displayed hippocampal neuron loss in patterns and extents similar but not identical to those reported previously for human patients with temporal lobe epilepsy. Similar to human patients, hippocampal sclerosis in sea lions was unilateral in 79% of cases, mossy fiber sprouting was a common neuropathological abnormality, and somatostatin-immunoreactive axons were exuberant in the dentate gyrus despite loss of immunopositive hilar neurons. Thus, hippocampal neuropathology of chronic DA sea lions is similar to that of human patients with temporal lobe epilepsy.”
Behavioral abnormalities and circuit defects in the Basal Ganglia of a mouse model of 16p11.2 deletion syndrome.
Thomas Portmann, Mu Yang, Rong Mao, Georgia Panagiotakos, Jacob Ellegood, Gul Dolen, Patrick Bader, Brad Grueter, Carleton Goold, Elaine Fisher, Katherine Clifford, Pavitra Rengarajan, David Kalikhman, Darren Loureiro, Nay Saw, Zhou Zhengqui, Michael Miller, Jason Lerch, R Mark Henkelman, Mehrdad Shamloo, Robert Malenka, Jacqueline Crawley, Ricardo Dolmetsch.
“A deletion on human chromosome 16p11.2 is associated with autism spectrum disorders. We deleted the syntenic region on mouse chromosome 7F3. MRI and high-throughput single-cell transcriptomics revealed anatomical and cellular abnormalities, particularly in cortex and striatum of juvenile mutant mice (16p11(+/-)). We found elevated numbers of striatal medium spiny neurons (MSNs) expressing the dopamine D2 receptor (Drd2(+)) and fewer dopamine-sensitive (Drd1(+)) neurons in deep layers of cortex. Electrophysiological recordings of Drd2(+) MSN revealed synaptic defects, suggesting abnormal basal ganglia circuitry function in 16p11(+/-) mice. This is further supported by behavioral experiments showing hyperactivity, circling, and deficits in movement control. Strikingly, 16p11(+/-) mice showed a complete lack of habituation reminiscent of what is observed in some autistic individuals. Our findings unveil a fundamental role of genes affected by the 16p11.2 deletion in establishing the basal ganglia circuitry and provide insights in the pathophysiology of autism.”
Extracellular architecture of the SYG-1/SYG-2 adhesion complex instructs synaptogenesis.
Engin Ozkan, Poh Hui Chia, Ruiqi Rachel Wang, Natalia Goriatcheva, Dominka Borek, Zbyszek Otwinowski, Thomas Walz, Kang Shen, K Christopher Garcia
“SYG-1 and SYG-2 are multipurpose cell adhesion molecules (CAMs) that have evolved across all major animal taxa to participate in diverse physiological functions, ranging from synapse formation to formation of the kidney filtration barrier. In the crystal structures of several SYG-1 and SYG-2 orthologs and their complexes, we find that SYG-1 orthologs homodimerize through a common, bispecific interface that similarly mediates an unusual orthogonal docking geometry in the heterophilic SYG-1/SYG-2 complex. C. elegans SYG-1’s specification of proper synapse formation in vivo closely correlates with the heterophilic complex affinity, which appears to be tuned for optimal function. Furthermore, replacement of the interacting domains of SYG-1 and SYG-2 with those from CAM complexes that assume alternative docking geometries or the introduction of segmental flexibility compromised synaptic function. These results suggest that SYG extracellular complexes do not simply act as “molecular velcro” and that their distinct structural features are important in instructing synaptogenesis.”
Visual space is compressed in prefrontal cortex before eye movements.
Marc Zirnsak, Nicholas Steinmetz, Behrad Noudoost, Kitty Xu, Tirin Moore
“We experience the visual world through a series of saccadic eye movements, each one shifting our gaze to bring objects of interest to the fovea for further processing. Although such movements lead to frequent and substantial displacements of the retinal image, these displacements go unnoticed. It is widely assumed that a primary mechanism underlying this apparent stability is an anticipatory shifting of visual receptive fields (RFs) from their presaccadic to their postsaccadic locations before movement onset. Evidence of this predictive ‘remapping’ of RFs has been particularly apparent within brain structures involved in gaze control. However, critically absent among that evidence are detailed measurements of visual RFs before movement onset. Here we show that during saccade preparation, rather than remap, RFs of neurons in a prefrontal gaze control area massively converge towards the saccadic target. We mapped the visual RFs of prefrontal neurons during stable fixation and immediately before the onset of eye movements, using multi-electrode recordings in monkeys. Following movements from an initial fixation point to a target, RFs remained stationary in retinocentric space. However, in the period immediately before movement onset, RFs shifted by as much as 18 degrees of visual angle, and converged towards the target location. This convergence resulted in a threefold increase in the proportion of RFs responding to stimuli near the target region. In addition, like in human observers, the population of prefrontal neurons grossly mislocalized presaccadic stimuli as being closer to the target. Our results show that RF shifts do not predict the retinal displacements due to saccades, but instead reflect the overriding perception of target space during eye movements.”
Gating of neural error signals during motor learning.
Rhea Kimpo*, Jacob Rinaldi*, Christina Kim*, Hannah Payne* and Jennifer Raymond
“Cerebellar climbing fiber activity encodes performance errors during many motor learning tasks, but the role of these error signals in learning has been controversial. We compared two motor learning paradigms that elicited equally robust putative error signals in the same climbing fibers: learned increases and decreases in the gain of the vestibulo-ocular reflex (VOR). During VOR-increase training, climbing fiber activity on one trial predicted changes in cerebellar output on the next trial, and optogenetic activation of climbing fibers to mimic their encoding of performance errors was sufficient to implant a motor memory. In contrast, during VOR-decrease training, there was no trial-by-trial correlation between climbing fiber activity and changes in cerebellar output, and climbing fiber activation did not induce VOR-decrease learning. Our data suggest that the ability of climbing fibers to induce plasticity can be dynamically gated in vivo, even under conditions where climbing fibers are robustly activated by performance errors. ”
Imaging neural spiking in brain tissue using FRET-opsin protein voltage sensors.
Yiyang Gond, Mark Wagner, Jin Zhong and Mark Schnitzer.
“Genetically encoded fluorescence voltage sensors offer the possibility of directly visualizing neural spiking dynamics in cells targeted by their genetic class or connectivity. Sensors of this class have generally suffered performance-limiting tradeoffs between modest brightness, sluggish kinetics and limited signalling dynamic range in response to action potentials. Here we describe sensors that use fluorescence resonance energy transfer (FRET) to combine the rapid kinetics and substantial voltage-dependence of rhodopsin family voltage-sensing domains with the brightness of genetically engineered protein fluorophores. These FRET-opsin sensors significantly improve upon the spike detection fidelity offered by the genetically encoded voltage sensor, Arclight, while offering faster kinetics and higher brightness. Using FRET-opsin sensors we imaged neural spiking and sub-threshold membrane voltage dynamics in cultured neurons and in pyramidal cells within neocortical tissue slices. In live mice, rates and optical waveforms of cerebellar Purkinje neurons’ dendritic voltage transients matched expectations for these cells’ dendritic spikes.”
Suppression of Alzheimer-associated inflammation by microglial prostaglandin-E2 EP4 receptor signaling.
Nathaniel Woodling, Qian Wang, Prachi Priyam, Paul Larkin, Ju Shi, Jenny Johansson, Irene Zagol-Ikapitte, Oliver Boutaud, Katrin Andreasson.
“A persistent and nonresolving inflammatory response to accumulating Aβ peptide species is a cardinal feature in the development of Alzheimer’s disease (AD). In response to accumulating Aβ peptide species, microglia, the innate immune cells of the brain, generate a toxic inflammatory response that accelerates synaptic and neuronal injury. Many proinflammatory signaling pathways are linked to progression of neurodegeneration. However, endogenous anti-inflammatory pathways capable of suppressing Aβ-induced inflammation represent a relatively unexplored area. Here we report that signaling through the prostaglandin-E2 (PGE2) EP4 receptor potently suppresses microglial inflammatory responses to Aβ42 peptides. In cultured microglial cells, EP4 stimulation attenuated levels of Aβ42-induced inflammatory factors and potentiated phagocytosis of Aβ42. Microarray analysis demonstrated that EP4 stimulation broadly opposed Aβ42-driven gene expression changes in microglia, with enrichment for targets of IRF1, IRF7, and NF-κB transcription factors. In vivo, conditional deletion of microglial EP4 in APPSwe-PS1ΔE9 (APP-PS1) mice conversely increased inflammatory gene expression, oxidative protein modification, and Aβ deposition in brain at early stages of pathology, but not at later stages, suggesting an early anti-inflammatory function of microglial EP4 signaling in the APP-PS1 model. Finally, EP4 receptor levels decreased significantly in human cortex with progression from normal to AD states, suggesting that early loss of this beneficial signaling system in preclinical AD development may contribute to subsequent progression of pathology.”
Neuronal activity promotes oligodendrogenesis and adaptive myelination in the mammalian brain.
Erin Gibson, David Purger, Chrisopher Mount, Andrea Goldstein, Grant Lin, Lauren Wood, Ingrid Inema, Sarah Miller, Gregor Bieri, J. Bradley Zuchero, Ben Barres, Pamelyn Woo, Hannes Vogel, Michelle Monje.
“Myelination of the central nervous system requires the generation of functionally mature oligodendrocytes from oligodendrocyte precursor cells (OPCs). Electrically active neurons may influence OPC function and selectively instruct myelination of an active neural circuit. In this work, we use optogenetic stimulation of the premotor cortex in awake, behaving mice to demonstrate that neuronal activity elicits a mitogenic response of neural progenitor cells and OPCs, promotes oligodendrogenesis, and increases myelination within the deep layers of the premotor cortex and subcortical white matter. We further show that this neuronal activity-regulated oligodendrogenesis and myelination is associated with improved motor function of the corresponding limb. Oligodendrogenesis and myelination appear necessary for the observed functional improvement, as epigenetic blockade of oligodendrocyte differentiation and myelin changes prevents the activity-regulated behavioral improvement.”
Long-term cognitive impairments and pathological alterations in a mouse model of repetitive mild traumatic brain injury.
Jian Luo, Andy Nguyen, Sual Villeda, Hui Zhang, Zhaoqing Ding, Derek Lindsey, Gregor Bieri, Joseph Castellano, Gary Beaupre, Tony Wyss-Coray.
“Mild traumatic brain injury (mTBI, also referred to as concussion) accounts for the majority of all traumatic brain injuries. The consequences of repetitive mTBI have become of particular concern for individuals engaged in certain sports or in military operations. Many mTBI patients suffer long-lasting neurobehavioral impairments. In order to expedite pre-clinical research and therapy development, there is a need for animal models that reflect the long-term cognitive and pathological features seen in patients. In the present study, we developed and characterized a mouse model of repetitive mTBI, induced onto the closed head over the left frontal hemisphere with an electromagnetic stereotaxic impact device. Using GFAP-luciferase bioluminescence reporter mice that provide a readout of astrocyte activation, we observed an increase in bioluminescence relative to the force delivered by the impactor after single impact and cumulative effects of repetitive mTBI. Using the injury parameters established in the reporter mice, we induced a repetitive mTBI in wild-type C57BL/6J mice and characterized the long-term outcome. Animals received repetitive mTBI showed a significant impairment in spatial learning and memory when tested at 2 and 6 months after injury. A robust astrogliosis and increased p-Tau immunoreactivity were observed upon post-mortem pathological examinations. These findings are consistent with the deficits and pathology associated with mTBI in humans and support the use of this model to evaluate potential therapeutic approaches.”
