Emilio Geijo-Barrientos

The effects of dopamine on the subthreshold electrophysiological responses of rat prefrontal cortex neurons in vitro.

The rat prefrontal cortex is densely innervated by dopaminergic fibres originating in the mesencephalic ventral tegmental area, and dopamine application in vivo has an inhibitory effect. We have studied the effects of dopamine on the persistent sodium current that is present in prefrontal cortex neurons and on the subthreshold electrophysiological responses generated by that current: a slow depolarization and a fast oscillatory activity. Experiments were made in coronal slices of rat frontal cortex (300–400 pm thickness) and intracellular recordings from regularly spiking cells were obtained with 3 M potassium acetate‐filled glass microelectrodes (80–150 MΩ). Dopamine was applied dissolved in the extracellular medium and, in current‐clamp recordings, reversibly inhibited the slow subthreshold depolarization. Dopamine was ineffective when applied after tetrodotoxin (1 μM) had blocked the action potentials. This inhibition was dose‐dependent in the range of 0.1–10 μM. Dopamine, applied at 10 μM, decreased the steady‐state firing frequency and also inhibited the subthreshold fast oscillatory activity. The currents activated in the subthreshold range were recorded with the single‐electrode voltage‐clamp technique and a clear persistent, tetrodotoxin‐sensitive component was isolated. This component was inhibited by 50% in a reversible way by 20 μM dopamine. These results show that dopamine increases the threshold for spike firing and suggest a mechanism for the inhibitory action of this neurotransmitter in the prefrontal cortex.

Variations in genes regulating neuronal migration predict reduced prefrontal cognition in schizophrenia and bipolar subjects from mediterranean Spain: A preliminary study

Both neural development and prefrontal cortex function are known to be abnormal in schizophrenia and bipolar disorder. In order to test the hypothesis that these features may be related with genes that regulate neuronal migration, we analyzed two genomic regions: the lissencephaly critical region (chromosome 17p) encompassing the LIS1 gene and which is involved in human lissencephaly; and the genes related to the platelet-activating-factor, functionally related to LIS1, in 52 schizophrenic patients, 36 bipolar I patients and 65 normal control subjects. In addition, all patients and the 25 control subjects completed a neuropsychological battery. Thirteen (14.8%) patients showed genetic variations in either two markers related with lissencephaly or in the platelet-activating-factor receptor gene. These patients performed significantly worse in the Wisconsin Card Sorting Test-Perseverative Errors in comparison with patients with no lissencephaly critical region/platelet-activating-factor receptor variations. The presence of lissencephaly critical region/platelet-activating-factor receptor variations was parametrically related to perseverative errors, and this accounted for 17% of the variance (P = 0.0001). Finally, logistic regression showed that poor Wisconsin Card Sorting Test-Perseverative Errors performance was the only predictor of belonging to the positive lissencephaly critical region/platelet-activating-factor receptor group. These preliminary findings suggest that the variations in genes involved in neuronal migration predict the severity of the prefrontal cognitive deficits in both disorders.

Role of voltage‐dependent calcium channels in stimulus–secretion coupling in rabbit carotid body chemoreceptor cells

We have defined Ca2+ channel subtypes expressed in rabbit carotid body (CB) chemoreceptor cells and their participation in the stimulus‐evoked catecholamine (CA) release. Ca2+ currents (ICa) activated at –30 mV, peaked at +10 mV and were fully blocked by 200 μm Cd2+. L‐type channels (sensitive to 2 μm nisoldipine) activated at –30 mV and carried 21 ± 2% of total ICa. Non‐L‐type channels activated at potentials positive to –10 mV and carried: N channels (sensitive to 1 μmω‐conotoxin‐GVIA) 16 ± 1% of total ICa, P/Q channels (sensitive to 3 μmω‐conotoxin‐MVIIC after nisoldipine plus GVIA) 23 ± 3% of total ICa and R channels (resistant to all blockers combined) 40 ± 3% of total ICa. CA release induced by hypoxia, hypercapnic acidosis, dinitrophenol (DNP) and high K+o in the intact CB was inhibited by 79–98% by 200 μm Cd2+. Hypoxia, hypercapnic acidosis and DNP, depolarized chemoreceptor cells and eventually generated repetitive action potential discharge. Nisoldipine plus MVIIC nearly abolished the release of CAs induced by hypoxia and hypercapnic acidosis and reduced by 74% that induced by DNP. All these secretory responses were insensitive to GVIA. 30 and 100 mm K+o brought resting membrane potential (Em) of chemoreceptor cells (–48.1 ± 1.2 mV) to –22.5 and +7.2 mV, respectively. Thirty millimolar K+o‐evoked release was abolished by nisoldipine but that induced by 100 mm K+o was mediated by activation of L, N, and P/Q channels. Data show that tested stimuli depolarize rabbit CB chemoreceptor cells and elicit CA release through Ca2+ entry via voltage‐activated channels. Only L and P/Q channels are tightly coupled to the secretion of CA.

CALCIUM-ACTIVATED CHLORIDE CURRENT IN NORMAL MOUSE SYMPATHETIC GANGLION CELLS

1. In rat sympathetic ganglion cells, axotomy induces the appearance of a depolarizing after‐potential (ADP) produced by a calcium‐activated chloride current. Here we report that this current is also present in normal sympathetic neurones from the mouse. 2. In an in vitro preparation of the superior cervical ganglion, an ADP was observed after spike firing in 50% of the cells studied with single‐electrode current‐ and voltage‐clamp techniques. 3. When the cells were voltage clamped at ‐50 mV in the presence of tetrodotoxin (TTX) and tetraethylammonium chloride (TEA), depolarizing jumps evoked inward calcium currents which were contaminated by outward chloride currents, followed by slowly decaying inward chloride tail currents. 4. The ADP and the inward tail currents disappeared when calcium was removed from the extracellular solution or when cadmium was added. 5. The reversal potential for the inward tail current was approximately ‐24 mV and was displaced in agreement with the Nernst equation for chloride when the extracellular NaCl was replaced by sucrose or sodium isethionate. The chloride channel blocker anthracene‐9‐carboxylic acid (9AC) inhibited both the ADP and the tail current. 6. Using intracellular injection of neurobiotin, we found that cells with shorter dendrites had larger ADPs. In axotomized ganglia practically all cells showed very pronounced ADPs. 7. We conclude that normal mouse sympathetic ganglion cells have a calcium‐activated chloride current that generates an ADP. The channels responsible for this current are probably located in the dendrites.

Evidence for association between structural variants in lissencephaly-related genes and executive deficits in schizophrenia or bipolar patients from a Spanish isolate population

There is evidence for an association between structural variants in genes for lissencephaly, which are involved in neuronal migration, and prefrontal cognitive deficits in schizophrenia and bipolar patients. On the basis of these intriguing findings, we analyzed 16 markers located in the lissencephaly critical region (LCR in chromosome 17p13.3) in 124 schizophrenic, 56 bipolar, and 141 healthy individuals. All recruits were from a Spanish population isolate of Basque origin that is characterized by low genetic heterogeneity. In addition, we examined whether structural genomic variations in the LCR were associated with executive cognition. Twenty-three patients (12.8%), but none of the controls, showed structural variants (deletions and insertions) in either of two markers related with lissencephaly (D17S1566 on tumor suppressor gene TP53: tumor protein p53 and D17S22 on SMG6 gene: Smg-6 homolog, nonsense mediated mRNA decay factor– Caenorhabditis elegans). These patients performed significantly worse in the Wisconsin Card Sorting Test-Categories in comparison with patients without such variations in lissencephaly-related genes. The presence of structural variants was related to completed categories, and accounted for 10.7% of the variance (P=0.001). Finally, logistic regression showed that poor Wisconsin Card Sorting Test-Categories performance was the only predictor of belonging to the positive LCR variations group. These new findings provide further evidence for the association between some lissencephaly-related genes and both schizophrenia and bipolar disorder, and influence on frontal executive functioning.

A study of F‐waves in patients with unilateral lumbosacral radiculopathy

Background and purpose:  The F wave, a late response of low amplitude, is widely used in the study of peripheral nerve lesions, and its persistence and latencies are the main parameters that are usually considered. The analysis of repeater F‐waves, which are commonly observed in association with focal or generalized motor neuropathy, is not always performed as a standard electrodiagnostic protocol.

Subthreshold inward membrane currents in guinea-pig frontal cortex neurons

Current-clamp and single-electrode voltage-clamp recordings were used to study the inward currents activated in the subthreshold membrane potential range of cortical pyramidal neurons. The experiments were done on slices from guinea-pig frontal cortex and all recordings were obtained at a distance of 600-900 microm from the pial surface. In current-clamp recordings and from membrane potentials hyperpolarized to about -70 mV, the depolarization leading to spike firing was partially blocked by 1 microM tetrodotoxin, but not by calcium-free extracellular solution. The calcium-free solution only affected this depolarization when the membrane potential was held at a level more negative than -75 mV. Under voltage-clamp, an inward current was recorded between the resting membrane potential and the level of spike firing. This current was activated at about -60 mV and part of it was blocked by 1 microM tetrodotoxin; the remaining current was blocked by calcium-free extracellular solution. In five neurons both components were recorded and isolated in the same cell. The tetrodotoxin-sensitive component activated at close to -60 mV, was similar to the persistent sodium current (I(Na-p)). The Ca2+-sensitive component activated at close to -60 or -65 mV, was less voltage-dependent than I(Na-p). This component was similar to the low threshold calcium current (I(T)). These results suggest that the subthreshold depolarization which led to spike firing was dependent on I(Na-p) and I(T), I(Na-p) being the most important factor up to resting membrane potentials of -70 or -75 mV. A physiological role of this finding is revealed by the action of dopamine, which (at 10 microM) prevented the firing of action potentials from -60 mV, but not from -80 mV due to the inhibition of I(Na-p) and the lack of effect on I(T).

Blocking miRNA Biogenesis in Adult Forebrain Neurons Enhances Seizure Susceptibility, Fear Memory, and Food Intake by Increasing Neuronal Responsiveness

The RNase Dicer is essential for the maturation of most microRNAs, a molecular system that plays an essential role in fine-tuning gene expression. To gain molecular insight into the role of Dicer and the microRNA system in brain function, we conducted 2 complementary RNA-seq screens in the hippocampus of inducible forebrain-restricted Dicer1 mutants aimed at identifying the microRNAs primarily affected by Dicer loss and their targets, respectively. Functional genomics analyses predicted the main biological processes and phenotypes associated with impaired microRNA maturation, including categories related to microRNA biology, signal transduction, seizures, and synaptic transmission and plasticity. Consistent with these predictions, we found that, soon after recombination, Dicer-deficient mice exhibited an exaggerated seizure response, enhanced induction of immediate early genes in response to different stimuli, stronger and more stable fear memory, hyperphagia, and increased excitability of CA1 pyramidal neurons. In the long term, we also observed slow and progressive excitotoxic neurodegeneration. Overall, our results indicate that interfering with microRNA biogenesis causes an increase in neuronal responsiveness and disrupts homeostatic mechanisms that protect the neuron against overactivation, which may explain both the initial and late phenotypes associated with the loss of Dicer in excitatory neurons.

Participation of low‐threshold calcium spikes in excitatory synaptic transmission in guinea pig medial frontal cortex

We studied the activation of low‐threshold calcium spikes (LTS) by excitatory postsynaptic potentials in pyramidal neurons from guinea pig medial frontal cortex with intracellular recording. We used extracellular bicuculline and phaclofen and intracellular QX‐314 to block inhibitory synaptic potentials and sodium currents. Postsynaptic potentials were evoked by stimulation of layer I. We found that large (> 10–15 mV) excitatory synaptic potentials evoked from membrane potentials more negative than −75 mV were able to trigger LTS. The activation of LTS resulted in an increase of the rising slope or amplitude of the synaptic potentials depending on the size of the excitatory postsynaptic potential (EPSP). We used 100 μm NiCl2 to confirm the presence of LTS as part of the EPSPs. The N‐methyl‐d‐aspartate (NMDA) and non‐NMDA components of the excitatory synaptic potentials were isolated using (±)2‐amino‐5‐phosphonovaleric acid (APV; 50 μm) or 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX; 20 μm); both components could, independently, trigger an LTS. With recordings made with K+ acetate‐filled electrodes, we show that the activation of LTS was critical to allow excitatory synaptic potentials to reach the threshold of action potential firing; also, this amplification of synaptic responses produced the firing of more than a single action potential by the postsynaptic cell. These results demonstrate that in cortical pyramidal neurons the activation of low‐threshold calcium spikes results in the amplification of synaptic responses.

Postnatal alterations of the inhibitory synaptic responses recorded from cortical pyramidal neurons in the Lis1/sLis1 mutant mouse

Mutations in the mouse Lis1 gene produce severe alterations in the developing cortex. We have examined some electrophysiological responses of cortical pyramidal neurons during the early postnatal development of Lis/sLis1 mutant mice. In P7 and P30 Lis1/sLis1 neurons we detected a lower frequency and slower decay phase of mIPSCs, and at P30 the mIPSCs amplitude and the action potential duration were reduced. Zolpidem (an agonist of GABAA receptors containing the alpha1 subunit) neither modified the amplitude nor the decay time of mIPSCs at P7 in Lis1/sLis1 neurons, whereas it increased the decay time at P30. The levels of GABAA receptor alpha1 subunit mRNA were reduced in the Lis1/sLis1 brain at P7 and P30, whereas reduced levels of the corresponding protein were only found at P7. These results demonstrate the presence of functional alterations in the postnatal Lis1/sLis1 cortex and point to abnormalities in GABAA receptor subunit switching processes during postnatal development.