Ian G. Davison

In Vivo Light-Induced Activation of Neural Circuitry in Transgenic Mice Expressing Channelrhodopsin-2

Channelrhodopsin-2 (ChR2) is a light-gated, cation-selective ion channel isolated from the green algae Chlamydomonas reinhardtii. Here, we report the generation of transgenic mice that express a ChR2-YFP fusion protein in the CNS for in vivo activation and mapping of neural circuits. Using focal illumination of the cerebral cortex and olfactory bulb, we demonstrate a highly reproducible, light-dependent activation of neurons and precise control of firing frequency in vivo. To test the feasibility of mapping neural circuits, we exploited the circuitry formed between the olfactory bulb and the piriform cortex in anesthetized mice. In the olfactory bulb, individual mitral cells fired action potentials in response to light, and their firing rate was not influenced by costimulated glomeruli. However, in piriform cortex, the activity of target neurons increased as larger areas of the bulb were illuminated to recruit additional glomeruli. These results support a model of olfactory processing that is dependent upon mitral cell convergence and integration onto cortical cells. More broadly, these findings demonstrate a system for precise manipulation of neural activity in the intact mammalian brain with light and illustrate the use of ChR2 mice in exploring functional connectivity of complex neural circuits in vivo.

Myosin Vb Mobilizes Recycling Endosomes and AMPA Receptors for Postsynaptic Plasticity

Learning-related plasticity at excitatory synapses in the mammalian brain requires the trafficking of AMPA receptors and the growth of dendritic spines. However, the mechanisms that couple plasticity stimuli to the trafficking of postsynaptic cargo are poorly understood. Here we demonstrate that myosin Vb (MyoVb), a Ca2+-sensitive motor, conducts spine trafficking during long-term potentiation (LTP) of synaptic strength. Upon activation of NMDA receptors and corresponding Ca2+ influx, MyoVb associates with recycling endosomes (REs), triggering rapid spine recruitment of endosomes and local exocytosis in spines. Disruption of MyoVb or its interaction with the RE adaptor Rab11-FIP2 abolishes LTP-induced exocytosis from REs and prevents both AMPA receptor insertion and spine growth. Furthermore, induction of tight binding of MyoVb to actin using an acute chemical genetic strategy eradicates LTP in hippocampal slices. Thus, Ca2+-activated MyoVb captures and mobilizes REs for AMPA receptor insertion and spine growth, providing a mechanistic link between the induction and expression of postsynaptic plasticity.

Syntaxin-4 Defines a Domain for Activity-Dependent Exocytosis in Dendritic Spines

Changes in postsynaptic membrane composition underlie many forms of learning-related synaptic plasticity in the brain. At excitatory glutamatergic synapses, fusion of intracellular vesicles at or near the postsynaptic plasma membrane is critical for dendritic spine morphology, retrograde synaptic signaling, and long-term synaptic plasticity. Whereas the molecular machinery for exocytosis in presynaptic terminals has been defined in detail, little is known about the location, kinetics, regulation, or molecules involved in postsynaptic exocytosis. Here, we show that an exocytic domain adjacent to the postsynaptic density (PSD) enables fusion of large, AMPA receptor-containing recycling compartments during elevated synaptic activity. Exocytosis occurs at microdomains enriched in the plasma membrane t-SNARE syntaxin 4 (Stx4), and disruption of Stx4 impairs both spine exocytosis and long-term potentiation (LTP) at hippocampal synapses. Thus, Stx4 defines an exocytic zone that directs membrane fusion for postsynaptic plasticity, revealing a novel specialization for local membrane traffic in dendritic spines.

Sparse and Selective Odor Coding by Mitral/Tufted Neurons in the Main Olfactory Bulb

The mammalian olfactory system recognizes an enormous variety of odorants carrying a wide range of important behavioral cues. In the main olfactory bulb (MOB), odorants are ultimately represented through the action potential activity of mitral/tufted cells (M/Ts), whose selectivity and tuning to odorant molecules are therefore fundamental determinants of MOB sensory coding. However, the sheer number and diversity of discrete olfactory stimuli has been a major barrier to comprehensively evaluating M/T selectivity. To address this issue, we assessed M/T odorant responses in anesthetized mice to a 348-odorant panel widely and systematically distributed throughout chemical space, presented both individually and in mixtures at behaviorally relevant concentrations. We found that M/T activation by odorants was markedly selective, with neurons responding robustly, sensitively, and reliably to only a highly restricted subset of stimuli. Multiple odorants activating a single neuron commonly shared clear structural similarity, but M/T tuning also frequently extended beyond obviously defined chemical categories. Cells typically responded to effective compounds presented both individually and in mixtures, although firing rates evoked by mixtures typically showed partial suppression. Response selectivity was further confirmed in awake animals by chronic recordings of M/Ts. These data indicate that individual M/Ts encode specific odorant attributes shared by only a small fraction of compounds and imply that the MOB relays the collective molecular features of an odorant stimulus through a restricted set of M/Ts, each narrowly tuned to a particular stimulus characteristic.

Dopamine Inhibits Mitral/Tufted→ Granule Cell Synapses in the Frog Olfactory Bulb

Synaptic interactions between the dendrites of mitral/tufted (MT) and granule cells (GCs) in the olfactory bulb are important for the determination of spatiotemporal firing patterns of MTs, which form an odor representation passed to higher brain centers. These synapses are subject to modulation from several sources originating both within and outside the bulb. We show that dopamine, presumably released by TH-positive local interneurons, reduces synaptic transmission from MTs to GCs. MT neurons express D2-like receptors (D2Rs), and both dopamine and the D2 agonist quinpirole decrease EPSC amplitude at the MT→ GC synapse. D2R activation also increases paired pulse facilitation and decreases the frequency of action potential-independent spontaneous miniature EPSCs in GCs, consistent with an effect on MT glutamate release downstream from Ca2+ influx. Analysis of spike-evoked Ca2+ transients in MT lateral dendrites additionally shows that quinpirole reduces Ca2+ influx preferentially at distal locations, possibly by reducing dendritic excitability via increased transient K+ channel availability. When the OB is activated physiologically by using odor stimuli, blocking D2Rs increases the power of GABAA-dependent oscillations in the local field potential. This demonstrates a functional role for the dopaminergic circuit during normal odor-evoked responses and for the modulation of dendritic release and excitability in neuronal circuit function. Regulation of spike invasion of lateral dendrites by transient K+ currents also may provide a mechanism for local outputs of MTs to be controlled dynamically via other neuromodulators or by postsynaptic potentials.

Neural mechanisms of social learning in the female mouse

Social interactions are often powerful drivers of learning. In female mice, mating creates a long-lasting sensory memory for the pheromones of the stud male that alters neuroendocrine responses to his chemosignals for many weeks. The cellular and synaptic correlates of pheromonal learning, however, remain unclear. We examined local circuit changes in the accessory olfactory bulb (AOB) using targeted ex vivo recordings of mating-activated neurons tagged with a fluorescent reporter. Imprinting led to striking plasticity in the intrinsic membrane excitability of projection neurons (mitral cells, MCs) that dramatically curtailed their responsiveness, suggesting a novel cellular substrate for pheromonal learning. Plasticity was selectively expressed in the MC ensembles activated by the stud male, consistent with formation of memories for specific individuals. Finally, MC excitability gained atypical activity-dependence whose slow dynamics strongly attenuated firing on timescales of several minutes. This unusual form of AOB plasticity may act to filter sustained or repetitive sensory signals. DOI: http://dx.doi.org/10.7554/eLife.25421.001

Olfactory receptor accessory proteins play crucial roles in receptor function and gene choice

Each of the olfactory sensory neurons (OSNs) chooses to express a single G protein-coupled olfactory receptor (OR) from a pool of hundreds. Here, we show the receptor transporting protein (RTP) family members play a dual role in both normal OR trafficking and determining OR gene choice probabilities. Rtp1 and Rtp2 double knockout mice (RTP1,2DKO) show OR trafficking defects and decreased OSN activation. Surprisingly, we discovered a small subset of the ORs are expressed in larger numbers of OSNs despite the presence of fewer total OSNs in RTP1,2DKO. Unlike typical ORs, some overrepresented ORs show robust cell surface expression in heterologous cells without the co-expression of RTPs. We present a model in which developing OSNs exhibit unstable OR expression until they choose to express an OR that exits the ER or undergo cell death. Our study sheds light on the new link between OR protein trafficking and OR transcriptional regulation. DOI: http://dx.doi.org/10.7554/eLife.21895.001

Phase-gradient contrast in thick tissue with a scanning microscope

It is well known that the principle of reciprocity is valid for light traveling even through scattering or absorptive media. This principle has been used to establish an equivalence between conventional widefield microscopes and scanning microscopes. We make use of this principle to introduce a scanning version of oblique back-illumination microscopy, or sOBM. This technique provides sub-surface phase-gradient and amplitude images from unlabeled tissue, in an epi-detection geometry. That is, it may be applied to arbitrarily thick tissue. sOBM may be implemented as a simple, cost-effective add-on with any scanning microscope, requiring only the availability of an extra input channel in the microscope electronics. We demonstrate here its implementation in combination with two-photon excited fluorescence (TPEF) microscopy and with coherent anti-Stokes Raman scattering (CARS) microscopy, applied to brain or spinal cord tissue imaging. In both cases, sOBM provides information on tissue morphology complementary to TPEF or CARS contrast. This information is obtained simultaneously and is automatically co-registered. Finally, we show that sOBM can be operated at video rate.

Neuronal imaging with ultrahigh dynamic range multiphoton microscopy

Multiphoton microscopes are hampered by limited dynamic range, preventing weak sample features from being detected in the presence of strong features, or preventing the capture of unpredictable bursts in sample strength. We present a digital electronic add-on technique that vastly improves the dynamic range of a multiphoton microscope while limiting potential photodamage. The add-on provides real-time negative feedback to regulate the laser power delivered to the sample, and a log representation of the sample strength to accommodate ultrahigh dynamic range without loss of information. No microscope hardware modifications are required, making the technique readily compatible with commercial instruments. Benefits are shown in both structural and in-vivo functional mouse brain imaging applications.

Functional Plasticity of Odor Representations during Motherhood

Summary Motherhood is accompanied by new behaviors aimed at ensuring the wellbeing of the offspring. Olfaction plays a key role in guiding maternal behaviors during this transition. We studied functional changes in the main olfactory bulb (OB) of mothers in mice. Using in vivo two-photon calcium imaging, we studied the sensory representation of odors by mitral cells (MCs). We show that MC responses to monomolecular odors become sparser and weaker in mothers. In contrast, responses to biologically relevant odors are spared from sparsening or strengthen. MC responses to mixtures and to a range of concentrations suggest that these differences between odor responses cannot be accounted for by mixture suppressive effects or gain control mechanisms. In vitro whole-cell recordings show an increase in inhibitory synaptic drive onto MCs. The increase of inhibitory tone may contribute to the general decrease in responsiveness and concomitant enhanced representation of specific odors.