Medial entorhinal cortex commissural input regulates the activity of spatially and object-tuned cells contributing to episodic memory.

Extensive interhemispheric projections connect many homotopic brain regions, including the hippocampal formation, but little is known as to how information transfer affects the functions supported by the target area. Here, we studied whether the commissural projections connecting the medial entorhinal cortices contribute to spatial coding, object coding, and memory. We demonstrate that input from the contralateral medial entorhinal cortex targets all major cell types in the superficial medial entorhinal cortex, modulating their firing rate. Notably, a fraction of responsive cells displayed object tuning and exhibited a reduction in their firing rate upon the inhibition of commissural input. In line with this finding are behavioral results that revealed the contribution of commissural input to episodic-like memory retrieval.

Septal cholinergic input to CA2 hippocampal region controls social novelty discrimination via nicotinic receptor-mediated disinhibition.

Acetylcholine (ACh), released in the hippocampus from fibers originating in the medial septum/diagonal band of Broca (MSDB) complex, is crucial for learning and memory. The CA2 region of the hippocampus has received increasing attention in the context of social memory. However, the contribution of ACh to this process remains unclear. Here, we show that in mice, ACh controls social memory. Specifically, MSDB cholinergic neurons inhibition impairs social novelty discrimination, meaning the propensity of a mouse to interact with a novel rather than a familiar conspecific. This effect is mimicked by a selective antagonist of nicotinic AChRs delivered in CA2. Ex vivo recordings from hippocampal slices provide insight into the underlying mechanism, as activation of nAChRs by nicotine increases the excitatory drive to CA2 principal cells via disinhibition. In line with this observation, optogenetic activation of cholinergic neurons in MSDB increases the firing of CA2 principal cells in vivo. These results point to nAChRs as essential players in social novelty discrimination by controlling inhibition in the CA2 region.

Local and Distant Input Controlling Excitation in Layer II of the Medial Entorhinal Cortex.

Layer II (LII) of the medial entorhinal cortex (MEC) comprises grid cells that support spatial navigation. The firing pattern of grid cells might be explained by attractor dynamics in a network, which requires either direct excitatory connectivity between phase-specific grid cells or indirect coupling via interneurons. However, knowledge regarding local networks that support in vivo activity is incomplete. Here we identified essential components of LII networks in the MEC. We distinguished four types of excitatory neurons that exhibit cell-type-specific local excitatory and inhibitory connectivity. Furthermore, we found that LII neurons contribute to the excitation of contralateral neurons in the corresponding layer. Finally, we demonstrated that the medial septum controls excitation in the MEC via two subpopulations of long-range GABAergic neurons that target distinct interneurons in LII, thereby disinhibiting local circuits. We thus identified local connections that could support attractor dynamics and external inputs that likely govern excitation in LII.

Selective reduction of AMPA currents onto hippocampal interneurons impairs network oscillatory activity.

Reduction of excitatory currents onto GABAergic interneurons in the forebrain results in impaired spatial working memory and altered oscillatory network patterns in the hippocampus. Whether this phenotype is caused by an alteration in hippocampal interneurons is not known because most studies employed genetic manipulations affecting several brain regions. Here we performed viral injections in genetically modified mice to ablate the GluA4 subunit of the AMPA receptor in the hippocampus (GluA4(HC-/-) mice), thereby selectively reducing AMPA receptor-mediated currents onto a subgroup of hippocampal interneurons expressing GluA4. This regionally selective manipulation led to a strong spatial working memory deficit while leaving reference memory unaffected. Ripples (125-250 Hz) in the CA1 region of GluA4(HC-/-) mice had larger amplitude, slower frequency and reduced rate of occurrence. These changes were associated with an increased firing rate of pyramidal cells during ripples. The spatial selectivity of hippocampal pyramidal cells was comparable to that of controls in many respects when assessed during open field exploration and zigzag maze running. However, GluA4 ablation caused altered modulation of firing rate by theta oscillations in both interneurons and pyramidal cells. Moreover, the correlation between the theta firing phase of pyramidal cells and position was weaker in GluA4(HC-/-) mice. These results establish the involvement of AMPA receptor-mediated currents onto hippocampal interneurons for ripples and theta oscillations, and highlight potential cellular and network alterations that could account for the altered working memory performance.

Long-range-projecting GABAergic neurons modulate inhibition in hippocampus and entorhinal cortex.

The hippocampus and entorhinal cortex play a pivotal role in spatial learning and memory. The two forebrain regions are highly interconnected via excitatory pathways. Using optogenetic tools, we identified and characterized long-range γ-aminobutyric acid-releasing (GABAergic) neurons that provide a bidirectional hippocampal-entorhinal inhibitory connectivity and preferentially target GABAergic interneurons. Activation of long-range GABAergic axons enhances sub- and suprathreshold rhythmic theta activity of postsynaptic neurons in the target areas.

Gap junctions between interneurons are required for normal spatial coding in the hippocampus and short-term spatial memory.

Gap junctions containing connexin 36 electrically couple interneurons in many brain regions and synchronize their activity. We used connexin-36 knock-out mice (Cx36(-/-)) to study the importance of electrical coupling between interneurons for spatial coding in the hippocampus and for different forms of hippocampus-dependent spatial memory. Recordings in behaving mice revealed that the spatial selectivity of hippocampal pyramidal neurons was reduced and less stable in Cx36(-/-) mice. Altered network activity was reflected in slower theta oscillations in the mutants. Temporal coding, assessed by determining the presence and characteristics of theta phase precession, had different dynamics in Cx36(-/-) mice compared with controls. At the behavioral level, Cx36(-/-) mice displayed impaired short-term spatial memory but normal spatial reference memory. These results highlight the functional role of electrically coupled interneurons for spatial coding and cognition. Moreover, they suggest that the precise spatial selectivity of place cells is not essential for normal performance on spatial tasks assessing associative long-term memory.

NMDA receptor ablation on parvalbumin-positive interneurons impairs hippocampal synchrony, spatial representations, and working memory.

Activity of parvalbumin-positive hippocampal interneurons is critical for network synchronization but the receptors involved therein have remained largely unknown. Here we report network and behavioral deficits in mice with selective ablation of NMDA receptors in parvalbumin-positive interneurons (NR1(PVCre-/-)). Recordings of local field potentials and unitary neuronal activity in the hippocampal CA1 area revealed altered theta oscillations (5-10 Hz) in freely behaving NR1(PVCre-/-) mice. Moreover, in contrast to controls, in NR1(PVCre-/-) mice the remaining theta rhythm was abolished by the administration of atropine. Gamma oscillations (35-85 Hz) were increased and less modulated by the concurrent theta rhythm in the mutant. Positional firing of pyramidal cells in NR1(PVCre-/-) mice was less spatially and temporally precise. Finally, NR1(PVCre-/-) mice exhibited impaired spatial working as well as spatial short- and long-term recognition memory but showed no deficits in open field exploratory activity and spatial reference learning.

Hippocampal theta rhythm and its coupling with gamma oscillations require fast inhibition onto parvalbumin-positive interneurons.

Hippocampal theta (5-10 Hz) and gamma (35-85 Hz) oscillations depend on an inhibitory network of GABAergic interneurons. However, the lack of methods for direct and cell-type-specific interference with inhibition has prevented better insights that help link synaptic and cellular properties with network function. Here, we generated genetically modified mice (PV-Deltagamma(2)) in which synaptic inhibition was ablated in parvalbumin-positive (PV+) interneurons. Hippocampal local field potential and unit recordings in the CA1 area of freely behaving mice revealed that theta rhythm was strongly reduced in these mice. The characteristic coupling of theta and gamma oscillations was strongly altered in PV-Deltagamma(2) mice more than could be accounted for by the reduction in theta rhythm only. Surprisingly, gamma oscillations were not altered. These data indicate that synaptic inhibition onto PV+ interneurons is indispensable for theta- and its coupling to gamma oscillations but not for rhythmic gamma-activity in the hippocampus. Similar alterations in rhythmic activity were obtained in a computational hippocampal network model mimicking the genetic modification, suggesting that intrahippocampal networks might contribute to these effects.

Augmented hippocampal ripple oscillations in mice with reduced fast excitation onto parvalbumin-positive cells.

Generation of fast network oscillations in the hippocampus relies on interneurons, but the underlying specific synaptic mechanisms are not established. The excitatory recruitment of fast-spiking interneurons during hippocampal sharp waves has been suggested to be critical for the generation of 140-200 Hz ("ripple") oscillations in the CA1 area. To directly test this, we used genetically modified mice (PV-DeltaGluR-A) with reduced AMPA receptor-mediated excitation onto parvalbumin (PV)-positive interneurons and studied hippocampal oscillations in freely moving animals. In PV-DeltaGluR-A mice, ripple-amplitude and associated rhythmic modulation of pyramidal cells and fast-spiking interneurons were increased. These changes were not accompanied by concurrent alterations of firing rates. Neither theta nor gamma oscillations displayed marked alterations in the mutant. These results provide evidence that fast excitation from pyramidal cells to PV-positive interneurons differentially influences ripple and gamma oscillations in vivo.

Role of olivary electrical coupling in cerebellar motor learning.

The level of electrotonic coupling in the inferior olive is extremely high, but its functional role in cerebellar motor control remains elusive. Here, we subjected mice that lack olivary coupling to paradigms that require learning-dependent timing. Cx36-deficient mice showed impaired timing of both locomotion and eye-blink responses that were conditioned to a tone. The latencies of their olivary spike activities in response to the unconditioned stimulus were significantly more variable than those in wild-types. Whole-cell recordings of olivary neurons in vivo showed that these differences in spike timing result at least in part from altered interactions with their subthreshold oscillations. These results, combined with analyses of olivary activities in computer simulations at both the cellular and systems level, suggest that electrotonic coupling among olivary neurons by gap junctions is essential for proper timing of their action potentials and thereby for learning-dependent timing in cerebellar motor control.

Excitotoxicity in vitro by NR2A- and NR2B-containing NMDA receptors.

Excitotoxicity, exacerbating acute brain damage from brain trauma or stroke, is mediated in part by excessive Ca(2+)-influx from prolonged NMDA receptor activation. However, the contribution to excitotoxicity by each of the main NMDAR subtypes in glutamatergic forebrain neurons, the NR2A- and NR2B-types, has remained enigmatic. Here, we investigated this issue by use of pharmacological and genetic tools in cultured cortical neurons. In wild-type neurons the contribution of the NMDA receptor subtypes to excitotoxicity changed with the age of the cultures. The blockade of NR2B-containing NMDA receptors prevented NMDA-mediated toxicity in young cultures after 14days in vitro (DIV14), but both subtypes triggered excitotoxicity in older (DIV21) cultures. Notably, blocking either of the two subtypes failed to prevent NMDA-elicited cell death, indicating that the remaining subtype triggers cell demise. Intriguingly, a neuroprotective aspect of the NR2A subtype became apparent at submaximal NMDA concentration only at DIV21. The NR2A subtype mediated NMDA toxicity as well as partial protection only if it carried a functional C-terminal domain. Upon deletion of this domain in the NR2A subtype, excitotoxicity was mediated entirely via the NR2B subtype, both at DIV14 and DIV21. Our findings predict that successful therapeutic intervention in stroke based on currently available NMDA receptor subtype-selective blockers is unlikely.

Recruitment of parvalbumin-positive interneurons determines hippocampal function and associated behavior.

Perisomatic inhibition provided by a subgroup of GABAergic interneurons plays a critical role in timing the output of pyramidal cells. To test their contribution at the network and the behavioral level, we generated genetically modified mice in which the excitatory drive was selectively reduced either by the knockout of the GluR-D or by conditional ablation of the GluR-A subunit in parvalbumin-positive cells. Comparable cell type-specific reductions of AMPA-mediated currents were obtained. Kainate-induced gamma oscillations exhibited reduced power in hippocampal slices from GluR-D-/- and GluR-A(PVCre-/-) mice. Experimental and modeling data indicated that this alteration could be accounted for by imprecise spike timing of fast-spiking cells (FS) caused by smaller interneuronal EPSPs. GluR-D-/- and GluR-A(PVCre-/-) mice exhibited similar impairments in hippocampus-dependent tasks. These findings directly show the effects of insufficient recruitment of fast-spiking cells at the network and behavioral level and demonstrate the role of this subpopulation for working and episodic-like memory.

A reporter allele for investigating connexin 26 gene expression in the mouse brain.

A variety of connexins are expressed in the diverse cell types of the central nervous system and are thought to regulate some of the functional properties exhibited by immature and mature cells. A proper understanding of the role of specific connexins in these processes requires an unambiguous characterization of their spatial and temporal pattern of expression. In order to define the cellular distribution of connexin 26 (Cx26) in the mouse we have generated a reporter allele (Cx26lacZ) by genetically manipulating the locus so that the beta-galactosidase (lacZ) gene is expressed from the endogenous Cx26 promoter. This modification decreased expression from the allele and resulted in embryonic lethality for the Cx26lacZ/lacZ genotype in accordance with previous studies on Cx26 knock-out animals indicating that Cx26-containing gap junctions are necessary for embryonic development. Despite the lower than expected transcript levels, the amount of lacZ protein produced in heterozygous mice was sufficient to label tissues known to contain Cx26, such as liver, kidney, skin, cochlea, small intestine, placenta and thyroid gland. In the embryonic and mature central nervous system, however, lacZ was restricted to meningeal cells and could not be detected in either neurons or glia. The absence of Cx26 mRNA in these cells could also be confirmed by reverse transcription-polymerase chain reaction and in situ hybridization. Our experiments indicate that the Cx26lacZ mouse line can be used as a reporter of Cx26 gene expression and suggest that Cx26, contrary to previous reports, is restricted to the meninges in both embryonic and adult brain.