Paradoxical neuronal hyperexcitability in a mouse model of mitochondrial pyruvate import deficiency.

Neuronal excitation imposes a high demand of ATP in neurons. Most of the ATP derives primarily from pyruvate-mediated oxidative phosphorylation, a process that relies on import of pyruvate into mitochondria occuring exclusively via the mitochondrial pyruvate carrier (MPC). To investigate whether deficient oxidative phosphorylation impacts neuron excitability, we generated a mouse strain carrying a conditional deletion of MPC1, an essential subunit of the MPC, specifically in adult glutamatergic neurons. We found that, despite decreased levels of oxidative phosphorylation and decreased mitochondrial membrane potential in these excitatory neurons, mice were normal at rest. Surprisingly, in response to mild inhibition of GABA mediated synaptic activity, they rapidly developed severe seizures and died, whereas under similar conditions the behavior of control mice remained unchanged. We report that neurons with a deficient MPC were intrinsically hyperexcitable as a consequence of impaired calcium homeostasis, which reduced M-type potassium channel activity. Provision of ketone bodies restored energy status, calcium homeostasis and M-channel activity and attenuated seizures in animals fed a ketogenic diet. Our results provide an explanation for the seizures that frequently accompany a large number of neuropathologies, including cerebral ischemia and diverse mitochondriopathies, in which neurons experience an energy deficit.

A Translaminar Genetic Logic for the Circuit Identity of Intracortically Projecting Neurons.

Neurons of the neocortex are organized into six radial layers, which have appeared at different times during evolution, with the superficial layers representing a more recent acquisition. Input to the neocortex predominantly reaches superficial layers (SL, i.e., layers (L) 2-4), while output is generated in deep layers (DL, i.e., L5-6) [1]. Intracortical connections, which bridge input and output pathways, are key components of cortical circuits because they allow the propagation and processing of information within the neocortex. Two main types of intracortically projecting neurons (ICPN) can be distinguished by their axonal features: L4 spiny stellate neurons (SSN) with short axons projecting locally within cortical columns [2-5], and SL and DL long-range projection neurons, including callosally projecting neurons (CPNSL and CPNDL) [5, 6]. Here, we investigate the molecular hallmarks that distinguish SSN, CPNSL, and CPNDL and relate their transcriptional signatures with their output connectivity. Specifically, taking advantage of the presence of CPN in both SL and DL, we identify lamina-independent genetic hallmarks of a constant projection motif (i.e., interhemispheric projection). By performing unbiased transcriptomic comparisons between CPNSL, CPNDL and SSN, we provide specific molecular profiles for each of these populations and show that target identity supersedes laminar position in defining ICPN transcriptional diversity. Together, these findings reveal a projection-based organization of transcriptional programs across cortical layers, which we propose reflects conserved strategy to protect canonical circuit structure (and hence function) across a diverse range of neuroanatomies.

In vivo rapid gene delivery into postmitotic neocortical neurons using iontoporation.

This protocol describes a method for directing the expression of genes of interest into postmitotic neocortical neurons in vivo. Microinjection of a DNA plasmid-amphiphilic molecule mix into the neocortex followed by delivery of an ad hoc electric pulse protocol during the first few days of life in mice allows rapid, focal and efficient expression of genes in postmitotic neurons. Compared with other gene delivery techniques such as in utero electroporation and viral infection, this method allows rapid (12 h), focal (50-200 µm), mosaic-like (50 to several hundred neurons) targeting of postmitotic neurons within existing circuits. This 'iontoporation' protocol, which can be completed within ∼20 min per mouse, allows straightforward assessment of genetic constructs in postmitotic cortical neurons and subsequent genetic, histological and physiological investigations of gene function.

In vivo reprogramming of circuit connectivity in postmitotic neocortical neurons.

The molecular mechanisms that control how progenitors generate distinct subtypes of neurons, and how undifferentiated neurons acquire their specific identity during corticogenesis, are increasingly understood. However, whether postmitotic neurons can change their identity at late stages of differentiation remains unknown. To study this question, we developed an electrochemical in vivo gene delivery method to rapidly manipulate gene expression specifically in postmitotic neurons. Using this approach, we found that the molecular identity, morphology, physiology and functional input-output connectivity of layer 4 mouse spiny neurons could be specifically reprogrammed during the first postnatal week by ectopic expression of the layer 5B output neuron-specific transcription factor Fezf2. These findings reveal a high degree of plasticity in the identity of postmitotic neocortical neurons and provide a proof of principle for postnatal re-engineering of specific neural microcircuits in vivo.

Tumoral prostate shows different expression pattern of somatostatin receptor 2 (SSTR2) and phosphotyrosine phosphatase SHP-1 (PTPN6) according to tumor progression.

Prostate proliferation is dependent of androgens and many peptide hormones. Recent reports suggest that SSTR2 and SHP-1 were two fundamental components on antiproliferative effect of somatostatin. Many studies on SHP-1 revealed that the expression of this protein was diminished or abolished in several of the cancer cell lines and tissues examined. However, it is necessary to confront the cell lines data with real situation in cancer cases. Our studies have shown that epithelial expressions of both proteins, SHP-1 and SSTR2, in normal and benign hyperplasia are localized in the luminal side of duct and acinar cells. Also, SSTR2 is expressed in stromal cells. In malignant prostate tissue, SHP-1 was diminished in 28/45 cases or absent in 12/45 cases, whereas SSTR2 epithelial was diminished in 38/45 cases or lost in only 2/45 cases. The intensity of immunostained was highly negative correlated with Gleason grade for two proteins.