Veronica Egger

Mechanisms of Lateral Inhibition in the Olfactory Bulb: Efficiency and Modulation of Spike-Evoked Calcium Influx into Granule Cells

Granule cells are axonless local interneurons that mediate lateral inhibitory interactions between the principal neurons of the olfactory bulb via dendrodendritic reciprocal synapses. This unusual arrangement may give rise to functional properties different from conventional lateral inhibition. Although granule cells spike, little is known about the role of the action potential with respect to their synaptic output. To investigate the signals that underlie dendritic release in these cells, two-photon microscopy in rat brain slices was used to image calcium transients in granule cell dendrites and spines. Action potentials evoked calcium transients throughout the dendrites, with amplitudes increasing with distance from soma and attaining a plateau level within the external plexiform layer, the zone of granule cell synaptic output. Transient amplitudes were, on average, equal in size in spines and adjacent dendrites. Surprisingly, both spine and dendritic amplitudes were strongly dependent on membrane potential, decreasing with depolarization and increasing with hyperpolarization from rest. Both the current-voltage relationship and the time course of inactivation were consistent with the known properties of T-type calcium channels, and the voltage dependence was blocked by application of the T-type calcium channel antagonists Ni2+ and mibefradil. In addition, mibefradil reduced action potential-mediated synaptic transmission from granule to mitral cells. The implication of a transiently inactivating calcium channel in synaptic release from granule cells suggests novel mechanisms for the regulation of lateral inhibition in the olfactory bulb.

Synaptic inhibition in the olfactory bulb accelerates odor discrimination in mice.

Local inhibitory circuits are thought to shape neuronal information processing in the central nervous system, but it remains unclear how specific properties of inhibitory neuronal interactions translate into behavioral performance. In the olfactory bulb, inhibition of mitral/tufted cells via granule cells may contribute to odor discrimination behavior by refining neuronal representations of odors. Here we show that selective deletion of the AMPA receptor subunit GluA2 in granule cells boosted synaptic Ca(2+) influx, increasing inhibition of mitral cells. On a behavioral level, discrimination of similar odor mixtures was accelerated while leaving learning and memory unaffected. In contrast, selective removal of NMDA receptors in granule cells slowed discrimination of similar odors. Our results demonstrate that inhibition of mitral cells controlled by granule cell glutamate receptors results in fast and accurate discrimination of similar odors. Thus, spatiotemporally defined molecular perturbations of olfactory bulb granule cells directly link stimulus similarity, neuronal processing time, and discrimination behavior to synaptic inhibition.

Dendrodendritic Synaptic Signals in Olfactory Bulb Granule Cells: Local Spine Boost and Global Low-Threshold Spike

In the mammalian olfactory bulb, axonless granule cells process synaptic input and output reciprocally within large spines. The nature of the calcium signals that underlie the presynaptic and postsynaptic function of these spines is mostly unknown. Using two-photon imaging in acute rat brain slices and glomerular stimulation of mitral/tufted cells, we observed two forms of action potential-independent synaptic Ca2+ signals in granule cell dendrites. Weak activation of mitral/tufted cells produced stochastic Ca2+ transients in individual granule cell spines. These transients were strictly localized to the spine head, indicating a local passive boosting or spine spike. Ca2+ sources for these local synaptic events included NMDA receptors, voltage-dependent calcium channels, and Ca2+-induced Ca2+ release from internal stores. Stronger activation of mitral/tufted cells produced a low-threshold Ca2+ spike (LTS) throughout the granule cell apical dendrite. This global spike was mediated by T-type Ca2+ channels and represents a candidate mechanism for subthreshold lateral inhibition in the olfactory bulb. The coincidence of local input and LTS in the spine resulted in summation of local and global Ca2+ signals, a dendritic computation that could endow granule cells with subthreshold associative plasticity.

Local Postsynaptic Voltage-Gated Sodium Channel Activation in Dendritic Spines of Olfactory Bulb Granule Cells

Neuronal dendritic spines have been speculated to function as independent computational units, yet evidence for active electrical computation in spines is scarce. Here we show that strictly local voltage-gated sodium channel (Nav) activation can occur during excitatory postsynaptic potentials in the spines of olfactory bulb granule cells, which we mimic and detect via combined two-photon uncaging of glutamate and calcium imaging in conjunction with whole-cell recordings. We find that local Nav activation boosts calcium entry into spines through high-voltage-activated calcium channels and accelerates postsynaptic somatic depolarization, without affecting NMDA receptor-mediated signaling. Hence, Nav-mediated boosting promotes rapid output from the reciprocal granule cell spine onto the lateral mitral cell dendrite and thus can speed up recurrent inhibition. This striking example of electrical compartmentalization both adds to the understanding of olfactory network processing and broadens the general view of spine function.

Calcium buffering in rodent olfactory bulb granule cells and mitral cells

In the mammalian olfactory bulb, axonless granule cells (GCs) mediate self‐ and lateral inhibitory interactions between mitral cells (MCs) via reciprocal dendrodendritic synapses. Calcium signals in the GC dendrites and reciprocal spines appear to decay unusually slowly, hence GC calcium handling might contribute to the known asynchronous release at this synapse. By recording fluorescence transients of different Ca2+‐sensitive dyes at variable concentrations evoked by backpropagating action potentials (APs) and saturating AP trains we extrapolated Ca2+ dynamics to conditions of zero added buffer for juvenile rat GC apical dendrites and spines and MC lateral dendrites. Resting [Ca2+] was at ∼50 nm in both GC dendrites and spines. The average endogenous GC buffer capacities (κE) were within a range of 80–90 in the dendrites and 110–140 in the spines. The extrusion rate (γ) was estimated as 570 s−1 for dendrites and 870 s−1 for spines and the decay time constant as ∼200 ms for both. Single‐current‐evoked APs resulted in a [Ca2+] elevation of ∼250 nm. Calcium handling in juvenile and adult mouse GCs appeared mostly similar. In MC lateral dendrites, we found AP‐mediated [Ca2+] elevations of ∼130 nm with a similar decay to that in GC dendrites, while κE and γ were roughly 4‐fold higher. In conclusion, the slow GC Ca2+ dynamics are due mostly to sluggish Ca2+ extrusion. Under physiological conditions this slow removal may well contribute to delayed release and also feed into other Ca2+‐dependent mechanisms that foster asynchronous output from the reciprocal spine.

MRZ-99030 – a novel modulator of Aβ aggregation: II – reversal of Aβ oligomer-induced deficits in long-term potentiation (LTP) and cognitive performance in rats and mice.

β-amyloid1-42 (Aβ1-42) is a major endogenous pathogen underlying the aetiology of Alzheimers disease (AD). Recent evidence indicates that soluble Aβ oligomers, rather than plaques, are the major cause of synaptic dysfunction and neurodegeneration. Small molecules that suppress Aβ aggregation, reduce oligomer stability or promote off-pathway non-toxic oligomerization represent a promising alternative strategy for neuroprotection in AD. MRZ-99030 was recently identified as a dipeptide that modulates Aβ1-42 aggregation by triggering a non-amyloidogenic aggregation pathway, thereby reducing the amount of intermediate toxic soluble oligomeric Aβ species. The present study evaluated the relevance of these promising results with MRZ-99030 under pathophysiological conditions i.e. against the synaptotoxic effects of Aβ oligomers on hippocampal long term potentiation (LTP) and two different memory tasks. Aβ1-42 interferes with the glutamatergic system and with neuronal Ca(2+) signalling and abolishes the induction of LTP. Here we demonstrate that MRZ-99030 (100-500 nM) at a 10:1 stoichiometric excess to Aβ clearly reversed the synaptotoxic effects of Aβ1-42 oligomers on CA1-LTP in murine hippocampal slices. Co-application of MRZ-99030 also prevented the two-fold increase in resting Ca(2+) levels in pyramidal neuron dendrites and spines triggered by Aβ1-42 oligomers. In anaesthetized rats, pre-administration of MRZ-99030 (50 mg/kg s.c.) protected against deficits in hippocampal LTP following i.c.v. injection of oligomeric Aβ1-42. Furthermore, similar treatment significantly ameliorated cognitive deficits in an object recognition task and under an alternating lever cyclic ratio schedule after the i.c.v. application of Aβ1-42 and 7PA2 conditioned medium, respectively. Altogether, these results demonstrate the potential therapeutic benefit of MRZ-99030 in AD.

Dendritic Arborization Patterns of Small Juxtaglomerular Cell Subtypes within the Rodent Olfactory Bulb

Within the glomerular layer of the rodent olfactory bulb, numerous subtypes of local interneurons contribute to early processing of incoming sensory information. Here we have investigated dopaminergic and other small local juxtaglomerular cells in rats and mice and characterized their dendritic arborization pattern with respect to individual glomeruli by fluorescent labeling via patching and reconstruction of dendrites and glomerular contours from two-photon imaging data. Dopaminergic neurons were identified in a transgenic mouse line where the expression of dopamine transporter (DAT) was labeled with GFP. Among the DAT+ cells we found a small short-axon cell (SAC) subtype featuring hitherto undescribed dendritic specializations. These densely ramifying structures clasped mostly around somata of other juxtaglomerular neurons, which were also small, non-dopaminergic and to a large extent non-GABAergic. Clasping SACs were observed also in wild-type mice and juvenile rats. In DAT+ SAC dendrites, single backpropagating action potentials evoked robust calcium entry throughout both clasping and non-clasping compartments. Besides clasping SACs, most other small neurons either corresponded to the classical periglomerular cell type (PGCs), which was never DAT+, or were undersized cells with a small dendritic tree and low excitability. Aside from the presence of clasps in SAC dendrites, many descriptors of dendritic morphology such as the number of dendrites and the extent of branching were not significantly different between clasping SACs and PGCs. However, a detailed morphometric analysis in relation to glomerular contours revealed that the dendrites of clasping SACs arborized mostly in the juxtaglomerular space and never entered more than one glomerulus (if at all), whereas most PGC dendrites were restricted to their parent glomerulus, similar to the apical tufts of mitral cells. These complementary arborization patterns might underlie a highly complementary functional connectivity. The morphometric approach may serve to differentiate also other subtypes of juxtaglomerular neurons, help to identify putative synaptic partners and thus to establish a more refined picture of glomerular network interactions during odor sensing.

Suppression of SNARE-dependent exocytosis in retinal glial cells and its effect on ischemia-induced neurodegeneration

Nervous tissue is characterized by a tight structural association between glial cells and neurons. It is well known that glial cells support neuronal functions, but their role under pathologic conditions is less well understood. Here, we addressed this question in vivo using an experimental model of retinal ischemia and transgenic mice for glia‐specific inhibition of soluble N‐ethylmaleimide‐sensitive factor attachment protein receptor (SNARE)‐dependent exocytosis. Transgene expression reduced glutamate, but not ATP release from single Müller cells, impaired glial volume regulation under normal conditions and reduced neuronal dysfunction and death in the inner retina during the early stages of ischemia. Our study reveals that the SNARE‐dependent exocytosis in glial cells contributes to neurotoxicity during ischemia in vivo and suggests glial exocytosis as a target for therapeutic approaches.

Two-Photon Na+ Imaging Reports Somatically Evoked Action Potentials in Rat Olfactory Bulb Mitral and Granule Cell Neurites

Dendrodendritic synaptic interactions are a hallmark of neuronal processing in the vertebrate olfactory bulb. Many classes of olfactory bulb neurons including the principal mitral cells (MCs) and the axonless granule cells (GCs) dispose of highly efficient propagation of action potentials (AP) within their dendrites, from where they can release transmitter onto each other. So far, backpropagation in GC dendrites has been investigated indirectly via Ca2+ imaging. Here, we used two-photon Na+ imaging to directly report opening of voltage-gated sodium channels due to AP propagation in both cell types. To this end, neurons in acute slices from juvenile rat bulbs were filled with 1 mM SBFI via whole-cell patch-clamp. Calibration of SBFI signals revealed that a change in fluorescence ΔF/F by 10% corresponded to a Δ[Na+]i of ∼22 mM. We then imaged proximal axon segments of MCs during somatically evoked APs (sAP). While single sAPs were detectable in ∼50% of axons, trains of 20 sAPs at 50 Hz always resulted in substantial ΔF/F of ∼15% (∼33 mM Δ[Na+]i). ΔF/F was significantly larger for 80 Hz vs. 50 Hz trains, and decayed with half-durations τ1/2 ∼0.6 s for both frequencies. In MC lateral dendrites, AP trains yielded small ΔF/F of ∼3% (∼7 mM Δ[Na+]i). In GC apical dendrites and adjacent spines, single sAPs were not detectable. Trains resulted in an average dendritic ΔF/F of 7% (16 mM Δ[Na+]i) with τ1/2 ∼1 s, similar for 50 and 80 Hz. Na+ transients were indistinguishable between large GC spines and their adjacent dendrites. Cell-wise analysis revealed two classes of GCs with the first showing a decrease in ΔF/F along the dendrite with distance from the soma and the second an increase. These classes clustered with morphological parameters. Simulations of Δ[Na+]i replicated these behaviors via negative and positive gradients in Na+ current density, assuming faithful AP backpropagation. Such specializations of dendritic excitability might confer specific temporal processing capabilities to bulbar principal cell-GC subnetworks. In conclusion, we show that Na+ imaging provides a valuable tool for characterizing AP invasion of MC axons and GC dendrites and spines.

Brainstem-mediated sniffing and respiratory modulation during odor stimulation.

The trigeminal and olfactory systems interact during sensory processing of odor. Here, we investigate odor-evoked modulations of brainstem respiratory networks in a decerebrated perfused brainstem preparation of rat with intact olfactory bulbs. Intranasal application of non-trigeminal odors (rose) did not evoke respiratory modulation in absence of cortico-limbic circuits. Conversely, trigeminal odors such as menthol or lavender evoked robust respiratory modulations via direct activation of preserved brainstem circuits. Trigeminal odors consistently triggered short phrenic nerve bursts (fictive sniff), and the strong trigeminal odor menthol also triggered a slowing of phrenic nerve frequency. Phrenic and vagal nerve recordings reveal that fictive sniffs transiently interrupted odor evoked tonic postinspiratory vagal discharge. This motor pattern is significantly different from normal (eupneic) respiratory activity. In conclusion, we show for the first time the direct involvement of brainstem circuits in primary odor processing to evoke protective sniffs and respiratory modulation in the complete absence of forebrain commands.