HIV-1 Tat causes cognitive deficits and selective loss of parvalbumin, somatostatin, and neuronal nitric oxide synthase expressing hippocampal CA1 interneuron subpopulations.

Memory deficits are characteristic of HIV-associated neurocognitive disorders (HAND) and co-occur with hippocampal pathology. The HIV-1 transactivator of transcription (Tat), a regulatory protein, plays a significant role in these events, but the cellular mechanisms involved are poorly understood. Within the hippocampus, diverse populations of interneurons form complex networks; even subtle disruptions can drastically alter synaptic output, resulting in behavioral dysfunction. We hypothesized that HIV-1 Tat would impair cognitive behavior and injure specific hippocampal interneuron subtypes. Male transgenic mice that inducibly expressed HIV-1 Tat (or non-expressing controls) were assessed for cognitive behavior or had hippocampal CA1 subregions evaluated via interneuron subpopulation markers. Tat exposure decreased spatial memory in a Barnes maze and mnemonic performance in a novel object recognition test. Tat reduced the percentage of neurons expressing neuronal nitric oxide synthase (nNOS) without neuropeptide Y immunoreactivity in the stratum pyramidale and the stratum radiatum, parvalbumin in the stratum pyramidale, and somatostatin in the stratum oriens, which are consistent with reductions in interneuron-specific interneuron type 3 (IS3), bistratified, and oriens-lacunosum-moleculare interneurons, respectively. The findings reveal that an interconnected ensemble of CA1 nNOS-expressing interneurons, the IS3 cells, as well as subpopulations of parvalbumin- and somatostatin-expressing interneurons are preferentially vulnerable to HIV-1 Tat. Importantly, the susceptible interneurons form a microcircuit thought to be involved in feedback inhibition of CA1 pyramidal cells and gating of CA1 pyramidal cell inputs. The identification of vulnerable CA1 hippocampal interneurons may provide novel insight into the basic mechanisms underlying key functional and neurobehavioral deficits associated with HAND.

De novo expression of parvalbumin in ependymal cells in response to brain injury promotes ependymal remodeling and wound repair.

The calcium-binding protein parvalbumin (PV) hallmarks subpopulations of interneurons in the murine brain. We serendipitously observed the de novo expression of PV in ependymal cells of the lateral ventricle wall following in vivo lesioning and brain slicing for the preparation of organotypic hippocampal slice cultures (OHSCs). In OHSCs, de novo PV-expression begins shortly after the onset of culturing, and the number of ependymal cells implicated in this process increases with time. PV-immunopositive ependymal cells aggregate and form compact cell clusters, which are characterized by lumen-formation and beating cilia. Scratches inflicted on such clusters with a sharp knife are rapidly closed. Exposure of OHSCs to NF-КB-inhibitors and to antioxidants reduces PV-expression in ependymal cells, thereby implicating injury-induced inflammation in this process. Indeed, in vivo stab injury enhances PV-expression in ependymal cells adjacent to the lesion, whereas neuraminidase denudation is without effect. PV-knock-out mice manifest an impaired wound-healing response to in vivo injury, and a reduced scratch-wound reparation capacity in OHSCs. Whole-transcriptome analysis of ependymal-cell clusters in OHSCs revealed down-regulation of genes involved in cytoskeletal rearrangement, cell motility and cell adhesion in PV-knock out mice as compared with wild-type mice. Our data indicate that the injury-triggered up-regulation of PV-expression is mediated by inflammatory cytokines, and promotes the motility and adhesion of ependymal cells, thereby contributing to leakage closure by the re-establishment of a continuous ependymal layer.

Parvalbumin deficiency and GABAergic dysfunction in mice lacking PGC-1alpha.

The transcriptional coactivator peroxisome proliferator-activated receptor gamma coactivator 1alpha (PGC-1alpha) is a master regulator of metabolism in peripheral tissues, and it has been proposed that PGC-1alpha plays a similar role in the brain. Recent evidence suggests that PGC-1alpha is concentrated in GABAergic interneurons, so we investigated whether male and female PGC-1alpha -/- mice exhibit abnormalities in interneuron gene expression and/or function. We found a striking reduction in the expression of the Ca(2+)-binding protein parvalbumin (PV), but not other GABAergic markers, throughout the cerebrum in PGC-1alpha +/- and -/- mice. Furthermore, PGC-1alpha overexpression in cell culture was sufficient to robustly induce PV expression. Consistent with a reduction in PV rather than a loss of PV-expressing interneurons, spontaneous synaptic inhibition was not altered in PGC-1alpha -/- mice. However, evoked synaptic responses displayed less paired-pulse depression and dramatic facilitation in response to repetitive stimulation at the gamma frequency. PV transcript expression was also significantly reduced in retina and heart of PGC-1alpha -/- animals, suggesting that PGC-1alpha is required for proper expression of PV in multiple tissues. Together these findings indicate that PGC-1alpha is a novel regulator of interneuron gene expression and function and a potential therapeutic target for neurological disorders associated with interneuron dysfunction.

Somatic calcium level reports integrated spiking activity of cerebellar interneurons in vitro and in vivo.

We examined the relationship between somatic Ca²âº signals and spiking activity of cerebellar molecular layer interneurons (MLIs) in adult mice. Using two-photon microscopy in conjunction with cell-attached recordings in slices, we show that in tonically firing MLIs loaded with high-affinity Ca²âº probes, Ca²âº-dependent fluorescence transients are absent. Spike-triggered averages of fluorescence traces for MLIs spiking at low rates revealed that the fluorescence change associated with an action potential is small (1% of the basal fluorescence). To uncover the relationship between intracellular Ca²âº concentration ([Ca²âº](i)) and firing rates, spikes were transiently silenced with puffs of the GABA(A) receptor agonist muscimol. [Ca²âº](i) relaxed toward basal levels following a single exponential whose amplitude correlated to the preceding spike frequency. The relaxation time constant was slow (2.5 s) and independent of the probe concentration. Data from parvalbumin (PV)-/- animals indicate that PV controls the amplitude and decay time of spike-triggered averages as well as the time course of [Ca²âº](i) relaxations following spike silencing. The [Ca²âº](i) signals were sensitive to the L-type Ca²âº channel blocker nimodipine and insensitive to ryanodine. In anesthetized mice, as in slices, fluorescence traces from most MLIs did not show spontaneous transients. They nonetheless responded to muscimol iontophoresis with relaxations similar to those obtained in vitro, suggesting a state of tonic firing with estimated spiking rates ranging from 2 to 30 Hz. Altogether, the [Ca²âº](i) signal appears to reflect the integral of the spiking activity in MLIs. We propose that the muscimol silencing strategy can be extended to other tonically spiking neurons with similar [Ca²âº](i) homeostasis.

Operant conditioning deficits and modified local field potential activities in parvalbumin-deficient mice.

Altered functioning of GABAergic interneurons expressing parvalbumin (PV) in the basal ganglia-thalamo-cortical circuit are likely to be involved in several human psychiatric disorders characterized by deficits in attention and sensory gating with dysfunctional decision-making behavior. However, the contribution of these interneurons in the ability to acquire demanding learning tasks remains unclear. Here, we combine an operant conditioning task with local field potentials simultaneously recorded in several nuclei involved in reward circuits of wild-type (WT) and PV-deficient (PVKO) mice, which are characterized by changes in firing activity of PV-expressing interneurons. In comparison with WT mice, PVKO animals presented significant deficits in the acquisition of the selected learning task. Recordings from prefrontal cortex, nucleus accumbens (NAc) and hippocampus showed significant decreases of the spectral power in beta and gamma bands in PVKO compared with WT mice particularly during the performance of the operant conditioning task. From the first to the last session, at all frequency bands the spectral power in NAc tended to increase in WT and to decrease in PVKO. Results indicate that PV deficiency impairs signaling necessary for instrumental learning and the recognition of natural rewards.

Compensatory regulation of Cav2.1 Ca2+ channels in cerebellar Purkinje neurons lacking parvalbumin and calbindin D-28k.

Ca(v)2.1 channels regulate Ca(2+) signaling and excitability of cerebellar Purkinje neurons. These channels undergo a dual feedback regulation by incoming Ca(2+) ions, Ca(2+)-dependent facilitation and inactivation. Endogenous Ca(2+)-buffering proteins, such as parvalbumin (PV) and calbindin D-28k (CB), are highly expressed in Purkinje neurons and therefore may influence Ca(v)2.1 regulation by Ca(2+). To test this, we compared Ca(v)2.1 properties in dissociated Purkinje neurons from wild-type (WT) mice and those lacking both PV and CB (PV/CB(-/-)). Unexpectedly, P-type currents in WT and PV/CB(-/-) neurons differed in a way that was inconsistent with a role of PV and CB in acute modulation of Ca(2+) feedback to Ca(v)2.1. Ca(v)2.1 currents in PV/CB(-/-) neurons exhibited increased voltage-dependent inactivation, which could be traced to decreased expression of the auxiliary Ca(v)beta(2a) subunit compared with WT neurons. Although Ca(v)2.1 channels are required for normal pacemaking of Purkinje neurons, spontaneous action potentials were not different in WT and PV/CB(-/-) neurons. Increased inactivation due to molecular switching of Ca(v)2.1 beta-subunits may preserve normal activity-dependent Ca(2+) signals in the absence of Ca(2+)-buffering proteins in PV/CB(-/-) Purkinje neurons.

Absence of parvalbumin increases mitochondria volume and branching of dendrites in inhibitory Pvalb neurons in vivo: a point of convergence of autism spectrum disorder (ASD) risk gene phenotypes.

BACKGROUND: In fast firing, parvalbumin (PV)-expressing (Pvalb) interneurons, PV acts as an intracellular Ca2+ signal modulator with slow-onset kinetics. In Purkinje cells of PV-/- mice, adaptive/homeostatic mechanisms lead to an increase in mitochondria, organelles equally capable of delayed Ca2+ sequestering/buffering. An inverse regulation of PV and mitochondria likewise operates in cell model systems in vitro including myotubes, epithelial cells, and oligodendrocyte-like cells overexpressing PV. Whether such opposite regulation pertains to all Pvalb neurons is currently unknown. In oligodendrocyte-like cells, PV additionally decreases growth and branching of processes in a cell-autonomous manner. METHODS: The in vivo effects of absence of PV were investigated in inhibitory Pvalb neurons expressing EGFP, present in the somatosensory and medial prefrontal cortex, striatum, thalamic reticular nucleus, hippocampal regions DG, CA3, and CA1 and cerebellum of mice either wild-type or knockout (PV-/-) for the Pvalb gene. Changes in Pvalb neuron morphology and PV concentrations were determined using immunofluorescence, followed by 3D-reconstruction and quantitative image analyses. RESULTS: PV deficiency led to an increase in mitochondria volume and density in the soma; the magnitude of the effect was positively correlated with the estimated PV concentrations in the various Pvalb neuron subpopulations in wild-type neurons. The increase in dendrite length and branching, as well as thickness of proximal dendrites of selected PV-/- Pvalb neurons is likely the result of the observed increased density and length of mitochondria in these PV-/- Pvalb neuron dendrites. The increased branching and soma size directly linked to the absence of PV is assumed to contribute to the increased volume of the neocortex present in juvenile PV-/- mice. The extended dendritic branching is in line with the hypothesis of local hyperconnectivity in autism spectrum disorder (ASD) and ASD mouse models including PV-/- mice, which display all ASD core symptoms and several comorbidities including cortical macrocephaly at juvenile age. CONCLUSION: PV is involved in most proposed mechanisms implicated in ASD etiology: alterations in Ca2+ signaling affecting E/I balance, changes in mitochondria structure/function, and increased dendritic length and branching, possibly resulting in local hyperconnectivity, all in a likely cell autonomous way.

The role of parvalbumin and calbindin D28k in experimental scrapie.

AIMS: Prion diseases are generally characterized by pronounced neuronal loss. In particular, a subpopulation of inhibitory neurones, characterized by the expression of the calcium-binding protein parvalbumin (PV), is selectively destroyed early in the course of human and experimental prion diseases. By contrast, nerve cells expressing calbindin D28 k (CB), another calcium-binding protein, as well as PV/CB coexpressing Purkinje cells, are well preserved. METHODS: To evaluate, if PV and CB may directly contribute to neuronal vulnerability or resistance against nerve cell death, respectively, we inoculated PV- and CB-deficient mice, and corresponding controls, with 139A scrapie and compared them with regard to incubation times and histological lesion profiles. RESULTS: While survival times were slightly but significantly diminished in CB-/-, but not PV-/- mice, scrapie lesion profiles did not differ between knockout mice and controls. There was a highly significant and selective loss of isolectin B(4)-decorated perineuronal nets (which specifically demarcate the extracellular matrix surrounding the 'PV-expressing' subpopulation of cortical interneurones) in scrapie inoculated PV+/+, as well as PV-/- mice. Purkinje cell numbers were not different in CB+/+ and CB-/- mice. CONCLUSIONS: Our results suggest that PV expression is a surrogate marker for neurones highly vulnerable in prion diseases, but that the death of these neurones is unrelated to PV expression and thus based on a still unknown pathomechanism. Further studies including the inoculation of mice ectopically (over)expressing CB are necessary to determine whether the shortened survival of CB-/- mice is indeed due to a neuroprotective effect of this molecule.

Deficits in parvalbumin and calbindin immunoreactive cells in the hippocampus of isolation reared rats.

Post-mortem studies have provided evidence for abnormalities of the gamma-aminobutyric acid (GABA)-ergic system in schizophrenia. The calcium-binding proteins (CBPs), parvalbumin (PV), calbindin (CB) and calretinin (CR) can be used as markers for specific subpopulations of GABAergic neurons in the brain. Isolation rearing of rats is a non-pharmacological, non-lesion manipulation that leads to deficits in prepulse inhibition of the startle reflex (PPI) and other behavioural and neurochemical alterations reminiscent of schizophrenia. Female rats were reared in social housing (groups of three) or singly for 11 weeks post weaning and PPI was measured. Brains were removed and hippocampal CBP- containing neurons determined following immunocytochemical staining. Compared to socially housed rats, isolated rats exhibited PPI deficits and reductions in PV and CB-immunoreactive cells in the hippocampus, with no significant change in CR. These findings demonstrate selective abnormalities of sub-populations of GABAergic interneurons in the hippocampus of isolation reared rats, which resemble the neuronal deficits seen in this region in schizophrenia.

Deficiency in parvalbumin, but not in calbindin D-28k upregulates mitochondrial volume and decreases smooth endoplasmic reticulum surface selectively in a peripheral, subplasmalemmal region in the soma of Purkinje cells.

The Ca(2+)-binding proteins parvalbumin (PV) and calbindin D-28k (CB) are key players in the intracellular Ca(2+)-buffering in specific cells including neurons and have profound effects on spatiotemporal aspects of Ca(2+) transients. The previously observed increase in mitochondrial volume density in fast-twitch muscle of PV-/- mice is viewed as a specific compensation mechanism to maintain Ca(2+) homeostasis. Since cerebellar Purkinje cells (PC) are characterized by high expression levels of the Ca(2+) buffers PV and CB, the question was raised, whether homeostatic mechanisms are induced in PC lacking these buffers. Mitochondrial volume density, i.e. relative mitochondrial mass was increased by 40% in the soma of PV-/- PC. Upregulation of mitochondrial volume density was not homogenous throughout the soma, but was selectively restricted to a peripheral region of 1.5 microm width underneath the plasma membrane. Accompanied was a decreased surface of subplasmalemmal smooth endoplasmic reticulum (sPL-sER) in a shell of 0.5 microm thickness underneath the plasma membrane. These alterations were specific for the absence of the "slow-onset" buffer PV, since in CB-/- mice neither changes in peripheral mitochondria nor in sPL-sER were observed. This implicates that the morphological alterations are aimed to specifically substitute the function of the slow buffer PV. We propose a novel concept that homeostatic mechanisms of components involved in Ca(2+) homeostasis do not always occur at the level of similar or closely related molecules. Rather the cell attempts to restore spatiotemporal aspects of Ca(2+) signals prevailing in the undisturbed (wildtype) situation by subtly fine tuning existing components involved in the regulation of Ca(2+) fluxes.

Reduction in parvalbumin expression not loss of the parvalbumin-expressing GABA interneuron subpopulation in genetic parvalbumin and shank mouse models of autism.

BACKGROUND: A reduction of the number of parvalbumin (PV)-immunoreactive (PV(+)) GABAergic interneurons or a decrease in PV immunoreactivity was reported in several mouse models of autism spectrum disorders (ASD). This includes Shank mutant mice, with SHANK being one of the most important gene families mutated in human ASD. Similar findings were obtained in heterozygous (PV+/-) mice for the Pvalb gene, which display a robust ASD-like phenotype. Here, we addressed the question whether the observed reduction in PV immunoreactivity was the result of a decrease in PV expression levels and/or loss of the PV-expressing GABA interneuron subpopulation hereafter called "Pvalb neurons". The two alternatives have important implications as they likely result in opposing effects on the excitation/inhibition balance, with decreased PV expression resulting in enhanced inhibition, but loss of the Pvalb neuron subpopulation in reduced inhibition. METHODS: Stereology was used to determine the number of Pvalb neurons in ASD-associated brain regions including the medial prefrontal cortex, somatosensory cortex and striatum of PV-/-, PV+/-, Shank1-/- and Shank3B-/- mice. As a second marker for the identification of Pvalb neurons, we used Vicia Villosa Agglutinin (VVA), a lectin recognizing the specific extracellular matrix enwrapping Pvalb neurons. PV protein and Pvalb mRNA levels were determined quantitatively by Western blot analyses and qRT-PCR, respectively. RESULTS: Our analyses of total cell numbers in different brain regions indicated that the observed "reduction of PV(+) neurons" was in all cases, i.e., in PV+/-, Shank1-/- and Shank3B-/- mice, due to a reduction in Pvalb mRNA and PV protein, without any indication of neuronal cell decrease/loss of Pvalb neurons evidenced by the unaltered numbers of VVA(+) neurons. CONCLUSIONS: Our findings suggest that the PV system might represent a convergent downstream endpoint for some forms of ASD, with the excitation/inhibition balance shifted towards enhanced inhibition due to the down-regulation of PV being a promising target for future pharmacological interventions. Testing whether approaches aimed at restoring normal PV protein expression levels and/or Pvalb neuron function might reverse ASD-relevant phenotypes in mice appears therefore warranted and may pave the way for novel therapeutic treatment strategies.

Consequence of parvalbumin deficiency in the mdx mouse: histological, biochemical and mechanical phenotype of a new double mutant.

We tested the hypothesis whether the mild dystrophy in mdx mice could result from the contribution of the cytosolic calcium buffer parvalbumin in maintaining a normal cytosolic [Ca2+]i, in spite of an increased passive Ca2+ influx. By crossing mdx mice with parvalbumin-deficient mice, double mutant mice, lacking both dystrophin and parvalbumin, were obtained. Though resting cytosolic [Ca2+]i and total calcium content were similar to that of mdx muscles, this new animal model presented a slightly more severe phenotype than the mdx mouse. Muscle pseudo-hypertrophy, the density of myotubes and of centronucleated fibres as well as the loss of IIB fibres were all increased in parvalbumin-deficient mdx mice. Many of these deficits were overcome in late adulthood, albeit fibrosis was clearly more pronounced than in mdx muscles. At 90 days, parvalbumin-deficient mdx mice showed higher levels of creatine phosphokinase and lower muscle strength, in vivo, than mdx mice. Isometric tension of isolated muscle was reduced, but the susceptibility to eccentric contraction was not increased. The slight aggravation of muscle dystrophy observed in mdx mice deprived of parvalbumin cannot explain the severity of the affection observed in xmd dogs and Duchenne dystrophy patients where parvalbumin is constitutively not expressed.

A developmental cell-type switch in cortical interneurons leads to a selective defect in cortical oscillations.

The cellular diversity of interneurons in the neocortex is thought to reflect subtype-specific roles of cortical inhibition. Here we ask whether perturbations to two subtypes--parvalbumin-positive (PV+) and somatostatin-positive (SST+) interneurons--can be compensated for with respect to their contributions to cortical development. We use a genetic cell fate switch to delete both PV+ and SST+ interneurons selectively in cortical layers 2-4 without numerically changing the total interneuron population. This manipulation is compensated for at the level of synaptic currents and receptive fields (RFs) in the somatosensory cortex. By contrast, we identify a deficit in inhibitory synchronization in vitro and a large reduction in cortical gamma oscillations in vivo. This reveals that, while the roles of inhibition in establishing cortical inhibitory/excitatory balance and RFs can be subserved by multiple interneuron subtypes, gamma oscillations depend on cellular properties that cannot be compensated for--likely, the fast signalling properties of PV+ interneurons.

Parvalbumin cell ablation of NMDA-R1 causes increased resting network excitability with associated social and self-care deficits.

NMDA-receptor (NMDAR) hypofunction is strongly implicated in the pathophysiology of schizophrenia. Several convergent lines of evidence suggest that net excitation propagated by impaired NMDAR signaling on GABAergic interneurons may be of particular interest in mediating several aspects of schizophrenia. However, it is unclear which behavioral domains are governed by a net increase of excitation and whether modulating downstream GABAergic signaling can reverse neural and thus behavioral deficits. The current study determines the selective contributions of NMDAR dysfunction on PV-containing interneurons to electrophysiological, cognitive, and negative-symptom-related behavioral phenotypes of schizophrenia using mice with a PVcre-NR1flox-driven ablation of NR1 on PV-containing interneurons. In addition, we assessed the efficacy of one agent that directly modulates GABAergic signaling (baclofen) and one agent that indirectly modifies NMDAR-mediated signaling through antagonism of mGluR5 receptors (2-methyl-6-(phenylethynyl) pyridine (MPEP)). The data indicate that loss of NMDAR function on PV interneurons impairs self-care and sociability while increasing N1 latency and baseline gamma power, and reducing induction and maintenance of long-term potentiation. Baclofen normalized baseline gamma power without corresponding effects on behavior. MPEP further increased N1 latency and reduced social behavior in PVcre/NR1+/+ mice. These two indices were negatively correlated before and following MPEP such that as N1 latency increases, sociability decreases. This finding suggests a predictive role for N1 latency with respect to social function. Although previous data suggest that MPEP may be beneficial for core features of autism spectrum disorders, current data suggest that such effects require intact function of NMDAR on PV interneurons.

Visual thalamocortical circuits in parvalbumin-deficient mice.

The dorsal lateral geniculate nucleus (dLGN) is considered as the visual gateway to the visual cortex (VC) and sends collaterals to the thalamic reticular nucleus (RTN) that in turn receives collaterals of the corticofugal feedback projections. At all levels of this thalamocortical circuit there are GABAergic neurons expressing the calcium-buffer parvalbumin (PV). The present study reports for the first time the analysis of in vivo extracellular electrophysiological recordings performed simultaneously in dLGN, RTN and VC of anesthetized wild-type (WT) and parvalbumin-deficient (PVKO) mice. The firing rates of VC and RTN cells were increased in PVKO during spontaneous activity as well as in the presence of a photic stimulation (strobe flash at 2.5Hz). Interestingly, dLGN cells in PVKO did not show significant changes in the rate of firing in comparison to WT. dLGN responses to the light flashes were characterized by ripples of inhibition and phasic excitation/rebound. We have analyzed the pattern of functional interactions between pairs of neighboring cells in VC, dLGN and RTN and across these areas in simultaneously recorded thalamocortical triplets, with one neuron from each area. We found that in PVKO the strength of the interactions tended to decrease locally, between neighboring cells, but tended to increase across the areas. The combination of these analyses provides new evidence on the important role played by PV-expression in regulating information processing in the central visual pathway suggesting that the ability to process information along parallel channels is decreased in the thalamocortical pathway of PV-deficient mice. This article is part of a Special Issue entitled Neural Coding 2012.

Ambient illumination toggles a neuronal circuit switch in the retina and visual perception at cone threshold.

VIDEO ABSTRACT: Gradual changes in the sensory environment can lead to abrupt changes in brain computations and perception. However, mechanistic understanding of the mediating microcircuits is missing. By sliding through light levels from starlight to daylight, we identify retinal ganglion cell types in the mouse that abruptly and reversibly switch the weighting of center and surround interactions in their receptive field around cone threshold. Two-photon-targeted recordings and genetic and viral tracing experiments revealed that the circuit element responsible for the switch is a large inhibitory neuron that provides direct inhibition to ganglion cells. Our experiments suggest that weak excitatory input via electrical synapses together with the spiking threshold in inhibitory cells act as a switch. We also reveal a switch-like component in the spatial integration properties of human vision at cone threshold. This work demonstrates that circuits in the retina can quickly and reversibly switch between two distinct states, implementing distinct perceptual regimes at different light levels.

Inverse regulation of the cytosolic Ca²âº buffer parvalbumin and mitochondrial volume in muscle cells via SIRT1/PGC-1α axis.

Skeletal muscles show a high plasticity to cope with various physiological demands. Different muscle types can be distinguished by the force, endurance, contraction/relaxation kinetics (fast-twitch vs. slow-twitch muscles), oxidative/glycolytic capacity, and also with respect to Ca²âº-signaling components. Changes in Ca²âº signaling and associated Ca²âº-dependent processes are thought to underlie the high adaptive capacity of muscle fibers. Here we investigated the consequences and the involved mechanisms caused by the ectopic expression of the Ca²âº-binding protein parvalbumin (PV) in C2C12 myotubes in vitro, and conversely, the effects caused by its absence in in fast-twitch muscles of parvalbumin null-mutant (PV⁻/⁻) mice in vivo. The absence of PV in fast-twitch muscle tibialis anterior (TA) resulted in an increase in the peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α) and of its positive regulator, the deacetylase sirtuin 1 (SIRT1). TA muscles from PV⁻/⁻ mice also have an increased mitochondrial volume. Mild ionophore treatment of control (PV-devoid) C2C12 myotubes causing a moderate elevation in [Ca²âº](c) resulted in an increase in mitochondrial volume, together with elevated PGC-1α and SIRT1 expression levels, whilst it increased PV expression levels in myotubes stably transfected with PV. In PV-expressing myotubes the mitochondrial volume, PGC-1α and SIRT1 were significantly lower than in control C2C12 myotubes already at basal conditions and application of ionophore had no effect on either one. SIRT1 activation causes a down-regulation of PV in transfected myotubes, whilst SIRT1 inhibition has the opposite effect. We conclude that PV expression and mitochondrial volume in muscle cells are inversely regulated via a SIRT1/PGC-1α signaling axis.

Nuclear factor-κB is involved in the phenotype loss of parvalbumin-interneurons in vitro.

The phenotype loss of parvalbumin-containing interneurons, characterized by decreased parvalbumin expression, has been observed in schizophrenic patients. Overproduction of intraneuronal reactive oxygen species leads to such a phenotype loss. Nuclear factor-κB (NF-κB) activation is both a target and a regulator of intracellular oxidative stress response, suggesting its involvement in the parvalbumin regulation. This study was carried out to investigate the role of the NF-κB activation in the ketamine-induced phenotype loss of parvalbumin-interneurons in vitro. Ketamine was applied to primary neuronal cultures to successfully evoke the production of increased reactive oxygen species and decreased parvalbumin expression in parvalbumin-interneurons, which was invalid in the presence of a NF-κB inhibitor, SN50 or Bay11-7082. These results suggest potential links among NF-κB activation, oxidative stress, and parvalbumin-interneurons in vitro.

Neuregulin 1 represses limbic epileptogenesis through ErbB4 in parvalbumin-expressing interneurons.

Epilepsy is a common and refractory neurological disorder, but the neuronal regulatory mechanisms of epileptogenesis remain largely unclear. Activity-dependent transcription of genes for neurotrophins such as brain-derived neurotrophic factor (BDNF) has been shown to promote epileptogenesis; however, little is known about factors that may act as intrinsic, homeostatic or counterbalancing mechanisms. Using rodent models, here we show that limbic seizure activity upregulated NRG1-ErbB4 signaling and that epileptogenesis was inhibited by infusing NRG1 intracerebrally but exacerbated by neutralizing endogenous NRG1 with soluble ErbB4 extracellular domain, by inhibiting ErbB4 activation or by deleting the Erbb4 gene. Furthermore, specific depletion of ErbB4 in parvalbumin-expressing interneurons abolished NRG1-mediated inhibition of epileptogenesis and promoted kindling progression, resulting in increased spontaneous seizures and exuberant mossy fiber sprouting. In contrast, depleting ErbB4 in CaMKIIα-positive pyramidal neurons had no effect. Thus, NRG1-induced activation of ErbB4 in parvalbumin-expressing inhibitory interneurons may serve as a critical endogenous negative-feedback mechanism to suppress limbic epileptogenesis.

Upregulated expression of oncomodulin, the beta isoform of parvalbumin, in perikarya and axons in the diencephalon of parvalbumin knockout mice.

The calcium-binding proteins parvalbumin, calbindin D-28k, calretinin and calcineurin are present in subsets of GABAergic gigantic calyciform presynaptic terminals of the reticular thalamic nucleus (RTN). Previously it was hypothesized that GABA and calcium-binding proteins including parvalbumin are not only colocalized in the same neuron subpopulation, but that GABA synthesis and parvalbumin expression could be also genetically regulated by a common mechanism. Moreover, parvalbumin expression levels could influence GABA synthesis. For this, we analyzed GABA immunoreactivity in RTN gigantic calyciform presynaptic terminals of parvalbumin-deficient (PV-/-) mice. With respect to GABA immunoreactivity we found no differences compared to wild-type animals. However, using a polyclonal parvalbumin antibody raised against full-length rat muscle parvalbumin on brain sections of PV-/- mice, we observed paradoxical parvalbumin immunoreactivity in partly varicose axons in the diencephalon, mainly in the lamina medullaris externa surrounding the thalamus. A detailed immunohistochemical, biochemical and molecular biological analysis revealed this immunoreactivity to be the result of an upregulation of oncomodulin (OM), the mammalian beta isoform of parvalbumin in PV-/- mice. In addition, OM was present in a sparse subpopulation of neurons in the thalamus and in the dentate gyrus. OM expression has not been observed before in neurons of the mammalian brain; its expression was restricted to outer hair cells in the organ of Corti. Our results indicate that the absence of parvalbumin has no major effect on the GABA-synthesizing system in RTN presynaptic terminals excluding a direct effect of parvalbumin on this regulation. However, a likely homeostatic mechanism is induced resulting in the upregulation of OM in selected axons and neuronal perikarya. Our results warrant further detailed investigations on the putative role of OM in the brain.