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Cre-driver mouse lines have been extensively used as genetic tools to target and manipulate genetically defined neuronal populations by expression of Cre recombinase under selected gene promoters. This approach has greatly advanced neuroscience but interpretations are hampered by the fact that most Cre-driver lines have not been thoroughly characterized. Thus, a phenotypic characterization is of major importance to reveal potential aberrant phenotypes prior to implementation and usage to selectively inactivate or induce transgene expression. Here, we present a biochemical and behavioural assessment of the dopaminergic system in hemizygous tyrosine hydroxylase ( TH)-Cre mice in comparison to wild-type ( WT) controls. Our data show that TH-Cre mice display preserved dopaminergic homeostasis with unaltered levels of TH and dopamine as well as unaffected dopamine turnover in striatum. TH-Cre mice also show preserved dopamine transporter expression and function supporting sustained dopaminergic transmission. In addition, TH-Cre mice demonstrate normal responses in basic behavioural paradigms related to dopaminergic signalling including locomotor activity, reward preference and anxiolytic behaviour. Our results suggest that TH-Cre mice represent a valid tool to study the dopamine system, though careful characterization must always be performed to prevent false interpretations following Cre-dependent transgene expression and manipulation of selected neuronal pathways. [ABSTRACT FROM AUTHOR]

We studied the properties of dopaminergic neurons in the substantia nigra pars compacta ( SNpc) and ventral tegmental area ( VTA) in mice expressing the enhanced green fluorescent protein ( eGFP) under the control of the tyrosine hydroxylase promoter ( TH- GFP). By using a practical map of cell positioning in distinct SNpc and VTA subregions in horizontal midbrain slices we saw that the spontaneous firing, membrane properties, cell body size and magnitude of the hyperpolarization-activated current ( Ih) in TH- GFP-positive neurons ( TH- GFP+) vary significantly among subregions, following a mediolateral gradient. Block of Ih with Zd7288 inhibited firing in the most lateral subregions, but had little effect in the intermediate/medial VTA. In addition, TH- GFP+ cells were excited by Met5-Enkephalin. Extracellular recordings from a large neuron number showed that all TH- GFP+ cells were inhibited by dopamine, suggesting that this is a reliable approach for identifying dopaminergic neurons in vitro. Simultaneous recordings from dopamine-sensitive and dopamine-insensitive neurons showed that dopamine-insensitive cells (putative non-dopaminergic neurons) are unaffected by Zd7288 but inhibited by Met5-Enkephalin. Under patch-clamp, dopamine generated a quantitatively similar outward current in most TH- GFP+ neurons, although medial VTA cells showed reduced dopamine sensitivity. Pargyline prolonged the dopamine current, whereas cocaine enhanced dopamine-mediated responses in both the SNpc and the VTA. Our work provides new insights into the variability in mouse midbrain dopaminergic neurons along the medial-lateral axis and points to the necessity of a combination of different electrophysiological and pharmacological approaches for reliably identifying these cells to distinguish them from non-dopaminergic neurons in the midbrain. [ABSTRACT FROM AUTHOR]

Blockade of dopamine ( DA) reuptake via the dopamine transporter ( DAT) is a primary mechanism identified as underlying the therapeutic actions of (±)-modafinil (modafinil) and its R-enantiomer, armodafinil. Herein, we explored the neurochemical and behavioral actions of modafinil to better characterize its psychostimulant profile. Swiss-Webster mice were implanted with microdialysis probes in the nucleus accumbens shell ( NAS) or core ( NAC) to evaluate changes in DA levels related to acute reinforcing actions of drugs of abuse. Additionally, subjective effects were studied in mice trained to discriminate 10 mg/kg cocaine (i.p.) from saline. Modafinil (17-300 mg/kg, i.p.) significantly increased NAS and NAC DA levels that at the highest doses reached ~300% at 1 h, and lasted > 6 h in duration. These elevated DA levels did not show statistically significant regional differences between the NAS and NAC. Modafinil produced cocaine-like subjective effects at 56-100 mg/kg when administered at 5 and 60 min before the start of the session, and enhanced cocaine effects when the two were administered in combination. Despite sharing subjective effects with cocaine, modafinil's psychostimulant profile was unique compared to that of cocaine and like compounds. Modafinil had lower potency and efficacy than cocaine in stimulating NAS DA. In addition, the cocaine-like subjective effects of modafinil were obtained at lower doses and earlier onset times than expected based on its dopaminergic effects. These studies suggest that although inhibition of DA reuptake may be a primary mechanism underlying modafinil's therapeutic actions, non DA-dependent actions may be playing a role in its psychostimulant profile. [ABSTRACT FROM AUTHOR]

Stroke is a devastating neurological disease and the leading cause of long-term disability, particularly in the elderly. Calcium/calmodulin-dependent protein kinase kinase β (CaMKK β) is a major kinase activated by elevated levels of intracellular calcium. Our previous findings in young mice have suggested that Ca MKK β is neuroprotective as KO mice had worse stroke outcomes. Because age is an important determinant of stroke outcome, we evaluated the functional role of Ca MKK β in stroke in aged mice. We used middle cerebral artery occlusion to induce stroke in aged wild-type ( WT) and Ca MKK β KO male mice. Lentiviral vectors carrying Ca MKK β ( LV-Ca MKK β) were used to overexpress Ca MKK β in the mouse brain. Baseline levels of Ca MKK β in the aged brain were significantly lower than those in young mice. LV-Ca MKK β treatment reduced infarcts and neurological deficits assessed 3 days after stroke. In chronic survival experiments, Ca MKK β KO mice showed increased tissue loss in the ipsilateral hemisphere 3 weeks after stroke. In addition, KO mice showed poorer functional recovery during the 3-week survival period, as measured by the rotarod test, corner test, locomotor activity assay, and novel object recognition test, compared with WT controls. The loss of blood-brain barrier proteins, inactivation of survival gene expression such as B-cell lymphoma 2 (Bcl-2) and an increase in inflammatory cytokines in the serum were observed after stroke with Ca MKK β inhibition. We demonstrate that Ca MKK β is neuroprotective in stroke in aged mice. Therefore, our data suggest that Ca MKK β may be a potential target for reducing long-term disability after stroke. [ABSTRACT FROM AUTHOR]

Primitive neural stem cells (pNSCs) are the earliest NSCs to appear in the developing forebrain. They persist into the adult forebrain where they can generate all cells in the neural lineage and therefore hold great potential for brain regeneration. Thus, pNSCs are an ideal population to target to promote endogenous NSC activation. pNSCs can be isolated from the periventricular region as leukaemia inhibitory factor-responsive cells, and comprise a rare population in the adult mouse brain. We hypothesized that the pup periventricular region gives rise to more clonal pNSC-derived neurospheres but that pup-derived pNSCs are otherwise comparable to adult-derived pNSCs, and can be used to identify selective markers and activators of endogenous pNSCs. We tested the self-renewal ability, differentiation capacity and gene expression profile of pup-derived pNSCs and found them each to be comparable to adult-derived pNSCs, including being GFAP−, nestinmid, Oct4+. Next, we used pup pNSCs to test pharmacological compounds to activate pNSCs to promote endogenous brain repair. We hypothesized that pNSCs could be activated by targeting the cell surface proteins C-Kit and ErbB2, which were enriched in pNSCs relative to definitive NSCs (dNSCs) in an in vitro screen. C-Kit and ErbB2 signalling inhibition had distinct effects on pNSCs and dNSCs in vitro, and when infused directly into the adult brain in vivo. Targeted activation of pNSCs with C-Kit and ErbB2 modulation is a valuable strategy to activate the earliest cell in the neural lineage to contribute to endogenous brain regeneration. [ABSTRACT FROM AUTHOR]

The basal ganglia have long been implicated in action initiation. Using three-dimensional motion capture, we quantified the effects of optogenetic stimulation of the striatonigral (direct) pathway on movement kinematics. We generated transgenic mice with channelrhodopsin-2 expression in striatal neurons that express the D1-like dopamine receptor. With optic fibres placed in the sensorimotor striatum, an area known to contain movement velocity-related single units, photo-stimulation reliably produced movements that could be precisely quantified with our motion capture programme. A single light pulse was sufficient to elicit movements with short latencies (<30 ms). Increasing stimulation frequency increased movement speed, with a highly linear relationship. These findings support the hypothesis that the sensorimotor striatum is part of a velocity controller that controls rate of change in body configurations. [ABSTRACT FROM AUTHOR]

Pain is an emotion and neuropathic pain symptoms are modulated by supraspinal structures such as the amygdala. The central nucleus of the amygdala is often called the 'nociceptive amygdala', but little is known about the role of the basolateral amygdala. Here, we monitored the mechanical nociceptive thresholds in a mouse model of neuropathic pain and infused modulators of the glutamate/ GABAergic transmission in the basolateral nucleus of the amygdala ( BLA) via chronically-implanted cannulas. We found that an N-methyl-D-aspartate-type glutamate receptor antagonist ( MK-801) exerted a potent antiallodynic effect, whereas a transient allodynia was induced after perfusion of bicuculline, a GABAA receptor antagonist. Potentiating GABAA receptor function using diazepam or etifoxine (a non-benzodiazepine anxiolytic) fully but transiently alleviated mechanical allodynia. Interestingly, the antiallodynic effect of etifoxine disappeared in animals that were incapable of producing 3α-steroids. Diazepam had a similar effect but of shorter duration. As indicated by patch-clamp recordings of BLA neurons, these effects were mediated by a potentiation of GABAA receptor-mediated synaptic transmission. Together with a presynaptic elevation of miniature inhibitory postsynaptic current frequency, the duration and amplitude of GABAA miniature inhibitory postsynaptic currents were also increased (postsynaptic effect). The analgesic contribution of endogenous neurosteroid seemed to be exclusively postsynaptic. This study highlights the importance of the BLA and the local inhibitory/excitatory neuronal network activity while setting the mechanical nociceptive threshold. Furthermore, it appears that promoting inhibition in this specific nucleus could fully alleviate pain symptoms. Therefore, the BLA could be a novel interesting target for the development of pharmacological or non-pharmacological therapies. [ABSTRACT FROM AUTHOR]