Richardson N. Leão

OLM interneurons differentially modulate CA3 and entorhinal inputs to hippocampal CA1 neurons

The vast diversity of GABAergic interneurons is believed to endow hippocampal microcircuits with the required flexibility for memory encoding and retrieval. However, dissection of the functional roles of defined interneuron types has been hampered by the lack of cell-specific tools. We identified a precise molecular marker for a population of hippocampal GABAergic interneurons known as oriens lacunosum-moleculare (OLM) cells. By combining transgenic mice and optogenetic tools, we found that OLM cells are important for gating the information flow in CA1, facilitating the transmission of intrahippocampal information (from CA3) while reducing the influence of extrahippocampal inputs (from the entorhinal cortex). Furthermore, we found that OLM cells were interconnected by gap junctions, received direct cholinergic inputs from subcortical afferents and accounted for the effect of nicotine on synaptic plasticity of the Schaffer collateral pathway. Our results suggest that acetylcholine acting through OLM cells can control the mnemonic processes executed by the hippocampus.

A model of the electrically excited human cochlear neuron. II. Influence of the three-dimensional cochlear structure on neural excitability

A simplified spiraled model of the human cochlea is developed from a cross sectional photograph. The potential distribution within this model cochlea is calculated with the finite element technique for an active scala tympani implant. The method in the companion article [Rattay et al., 2001] allows for simulation of the excitation process of selected elements of the cochlear nerve. The bony boundary has an insulating influence along every nerve fiber which shifts the stimulation condition from that of a homogeneous extracellular medium towards constant field stimulation: for a target neuron which is stimulated by a ring electrode positioned just below the peripheral end of the fiber the extracellular voltage profile is rather linear. About half of the cochlear neurons of a completely innervated cochlea are excited with monopolar stimulation at three-fold threshold intensity, whereas bipolar and especially quadrupolar stimulation focuses the excited region even for stronger stimuli. In contrast to single fiber experiments with cats, the long peripheral processes in human cochlear neurons cause first excitation in the periphery and, consequently, neurons with lost dendrite need higher stimuli.

Topographic Organization in the Auditory Brainstem of Juvenile Mice is Disrupted in Congenital Deafness

There is an orderly topographic arrangement of neurones within auditory brainstem nuclei based on sound frequency. Previous immunolabelling studies in the medial nucleus of the trapezoid body (MNTB) have suggested that there may be gradients of voltage‐gated currents underlying this tonotopic arrangement. Here, our electrophysiological and immunolabelling results demonstrate that underlying the tonotopic organization of the MNTB is a combination of medio‐lateral gradients of low‐and high‐threshold potassium currents and hyperpolarization‐activated cation currents. Our results also show that the intrinsic membrane properties of MNTB neurones produce a topographic gradient of time delays, which may be relevant to sound localization, following previous demonstrations of the importance of the timing of inhibitory input from the MNTB to the medial superior olive (MSO). Most importantly, we demonstrate that, in the MNTB of congenitally deaf mice, which exhibit no spontaneous auditory nerve activity, the normal tonotopic gradients of neuronal properties are absent. Our results suggest an underlying mechanism for the observed topographic gradient of neuronal firing properties in the MNTB, show that an intrinsic neuronal mechanism is responsible for generating a topographic gradient of time‐delays, and provide direct evidence that these gradients rely on spontaneous auditory nerve activity during development.

On high-frequency field oscillations (>100 Hz) and the spectral leakage of spiking activity.

Recent reports converge to the idea that high-frequency oscillations in local field potentials (LFPs) represent multiunit activity. In particular, the amplitude of LFP activity above 100 Hz—commonly referred to as “high-gamma” or “epsilon” band—was found to correlate with firing rate. However, other studies suggest the existence of true LFP oscillations at this frequency range that are different from the well established ripple oscillations. Using multisite recordings of the hippocampus of freely moving rats, we show here that high-frequency LFP oscillations can represent either the spectral leakage of spiking activity or a genuine rhythm, depending on recording location. Both spike-leaked, spurious activity and true fast oscillations couple to theta phase; however, the two phenomena can be clearly distinguished by other key features, such as preferred coupling phase and spectral signatures. Our results argue against the idea that all high-frequency LFP activity stems from spike contamination and suggest avoiding defining brain rhythms solely based on frequency range.

Reduced low-voltage activated K+ conductances and enhanced central excitability in a congenitally deaf (dn/dn) mouse.

We have investigated changes in the neuronal excitability of the auditory brainstem in a congenitally deaf mouse (deafness dn/dn). Whole cell patch recordings from principal neurones of the medial nucleus of the trapezoid body (MNTB) showed strikingly enhanced excitability in the deaf mice when compared to control CBA mice at 12–14 days postnatal. MNTB neurones in normal CBA mice showed the phenotypic single action potential response on depolarization in current clamp; however, recordings from CBA mice carrying the homozygous deafness mutation fired trains of action potentials on depolarization. We show here that these changes are associated with reduced functional expression of dendrotoxin‐sensitive Kv1 potassium channels. In contrast, no differences were found in voltage‐gated calcium currents between control and deaf mice. These results reveal that loss of hair cell function in the cochlea leads to changes in ion channel expression in the central nervous system and suggests that this deafness model will be an important tool in understanding central changes occurring in human congenital deafness and in exploring activity‐dependent regulation of ion channel expression.

A transgenic mouse line for molecular genetic analysis of excitatory glutamatergic neurons

Excitatory glutamatergic neurons are part of most of the neuronal circuits in the mammalian nervous system. We have used BAC-technology to generate a BAC-Vglut2::Cre mouse line where Cre expression is driven by the vesicular glutamate transporter 2 (Vglut2) promotor. This BAC-Vglut2::Cre mouse line showed specific expression of Cre in Vglut2 positive cells in the spinal cord with no ectopic expression in GABAergic or glycinergic neurons. This mouse line also showed specific Cre expression in Vglut2 positive structures in the brain such as thalamus, hypothalamus, superior colliculi, inferior colliculi and deep cerebellar nuclei together with nuclei in the midbrain and hindbrain. Cre-mediated recombination was restricted to Cre expressing cells in the spinal cord and brain and occurred as early as E 12.5. Known Vglut2 positive neurons showed normal electrophysiological properties in the BAC-Vglut2::Cre transgenic mice. Altogether, this BAC-Vglut2::Cre mouse line provides a valuable tool for molecular genetic analysis of excitatory neuronal populations throughout the mouse nervous system.

Activity-dependent regulation of synaptic strength and neuronal excitability in central auditory pathways

Neural activity plays an important role in regulating synaptic strength and neuronal membrane properties. Attempts to establish guiding rules for activity‐dependent neuronal changes have led to such concepts as homeostasis of cellular activity and Hebbian reinforcement of synaptic strength. However, it is clear that there are diverse effects resulting from activity changes, and that these changes depend on the experimental preparation, and the developmental stage of the neural circuits under study. In addition, most experimental evidence on activity‐dependent regulation comes from reduced preparations such as neuronal cultures. This review highlights recent results from studies of the intact mammalian auditory system, where changes in activity have been shown to produce alterations in synaptic and membrane properties at the level of individual neurons, and changes in network properties, including the formation of tonotopic maps.

Hyperpolarization‐activated (Ih) currents in auditory brainstem neurons of normal and congenitally deaf mice

We have investigated the membrane properties of brainstem auditory neurons in a mouse model of congenital deafness (dn/dn). Whole‐cell recordings were made from visualized neurons in slices of the medial nucleus of the trapezoid body (MNTB) and anteroventral cochlear nucleus (AVCN). We have recently demonstrated that MNTB neurons in deaf mice are more excitable than in normal mice, due in part to a reduced expression of low‐threshold potassium currents. In this study, we have examined the contribution of hyperpolarization‐activated (Ih) channels to the membrane properties of MNTB and AVCN neurons. Our results show that Ih is larger in MNTB neurons from deaf mice than in normal mice. In contrast, no significant differences were found in Ih or excitability between AVCN bushy cells from dn/dn and normal mice. Experimental evidence and neuronal modelling suggests that, in the MNTB of normal mice, a small contribution of Ih helps to reduce temporal summation of synaptic potentials. A larger Ih in neurons from deaf mice has a much greater effect in reducing temporal summation of synaptic potentials, counteracting to some extent the greater excitability of these cells. Our results provide further insight into the role of activity during development in regulating the membrane and firing properties of central neurons.

Altered sodium currents in auditory neurons of congenitally deaf mice

Sodium currents are essential for action potential generation and propagation in most excitable cells. Appropriate tuning of these currents can be modulated both developmentally and in response to activity. Here we use a mouse model of congenital deafness (dn/dn– asymptomatic deafness associated with hair cell degeneration) to investigate the effect of lack of activity in the expression of Na+ currents in neurons from the medial nucleus of the trapezoid body (MNTB). Patch‐clamp recordings show that at postnatal day (P) 14, both normal and deaf mice display a significant amount of persistent and resurgent Na+ currents. However, the persistent current is greater in deaf mice than in normal mice, and resurgent current kinetics are slower in deaf mice. At P7, resurgent currents are not present in either group. MNTB immunohistochemistry demonstrates that Nav1.1 subunits are expressed postsynaptically in both P14 normal and deaf mice, while postsynaptic Nav1.6 staining was only observed in deaf mice. Labelling of Nav1.6 subunits in different age groups revealed that at younger ages (P7), both normal and deaf mice express this protein. Nav1.6 staining was not observed in MNTB neurons of P28 normal mice, whereas it is maintained in deaf mice cells until much later (P28). At P7, none of the groups displayed resurgent currents (despite the detection of Nav1.6 subunits at this age group); this suggests that factors other than alpha subunits are important for modulating these currents in MNTB cells. Our results emphasize the importance of activity during development in regulating Na+ channels.

Glycinergic mIPSCs in mouse and rat brainstem auditory nuclei: modulation by ruthenium red and the role of calcium stores

Spontaneous miniature inhibitory postsynaptic currents (mIPSCs) recorded in central neurons are usually highly variable in amplitude due to many factors such as intrinsic postsynaptic channel fluctuations at each release site, site‐to‐site variability between release sites, electrotonic attenuation due to variable dendritic locations of synapses, and the possibility of synchronous multivesicular release. A detailed knowledge of these factors is essential for the interpretation of mIPSC amplitude distributions and mean quantal size. We have studied glycinergic mIPSCs in two auditory brainstem nuclei, the rat anteroventral cochlear nucleus (AVCN) and the mouse medial nucleus of the trapezoid body (MNTB). Our previous results have demonstrated the location of glycinergic synapses on these neurons to be somatic, thus avoiding electrotonic complications. Spontaneous glycinergic mIPSCs were recorded from AVCN and MNTB neurons in brainstem slices, in the presence of TTX to block action potentials, and 6‐cyano‐7‐nitroquinoxaline‐2, 3‐dione, (±)‐2‐amino‐5‐phosphonopentanoic acid and bicuculline to block glutamatergic and GABAergic synaptic currents. Ruthenium red (RuR), which was used to increase the frequency of mIPSCs, significantly changed the shape of most (90 %) mIPSC amplitude distributions by increasing the proportion of large‐amplitude mIPSCs. The possibility was investigated (following previous evidence at GABAergic synapses) that large‐amplitude glycinergic mIPSCs are due to synchronous multivesicular release initiated by presynaptic calcium sparks from ryanodine‐sensitive calcium stores. Interval analysis of mIPSCs indicated that the number of potentially undetected (asynchrony < 0.5 ms) multivesicular mIPSCs was low in comparison with the number of large‐amplitude mIPSCs. Ryanodine, thapsigargin and calcium‐free perfusate did not reduce the frequency of large‐amplitude mIPSCs (> 150 pA), arguing against a significant role for presynaptic calcium stores. Our results support previous evidence suggesting that RuR increases miniature postsynaptic current (mSC) frequency by a mechanism that does not involve presynaptic calcium stores. Our results also indicate that at glycinergic synapses in the AVCN and MNTB, site‐to‐site variability in mIPSC amplitude, rather than multivesicular release, is a major factor underlying the large range of amplitudes of glycinergic mIPSCs.