Glutamate-system defects behind psychiatric manifestations in a familial hemiplegic migraine type 2 disease-mutation mouse model.

Migraine is a complex brain disorder, and understanding the complexity of this prevalent disease could improve quality of life for millions of people. Familial Hemiplegic Migraine type 2 (FHM2) is a subtype of migraine with aura and co-morbidities like epilepsy/seizures, cognitive impairments and psychiatric manifestations, such as obsessive-compulsive disorder (OCD). FHM2 disease-mutations locate to the ATP1A2 gene encoding the astrocyte-located α2-isoform of the sodium-potassium pump (α2Na(+)/K(+)-ATPase). We show that knock-in mice heterozygous for the FHM2-associated G301R-mutation (α2(+/G301R)) phenocopy several FHM2-relevant disease traits e.g., by mimicking mood depression and OCD. In vitro studies showed impaired glutamate uptake in hippocampal mixed astrocyte-neuron cultures from α2(G301R/G301R) E17 embryonic mice, and moreover, induction of cortical spreading depression (CSD) resulted in reduced recovery in α2(+/G301R) male mice. Moreover, NMDA-type glutamate receptor antagonists or progestin-only treatment reverted specific α2(+/G301R) behavioral phenotypes. Our findings demonstrate that studies of an in vivo relevant FHM2 disease knock-in mouse model provide a link between the female sex hormone cycle and the glutamate system and a link to co-morbid psychiatric manifestations of FHM2.

Fluidic system for long-term in vitro culturing and monitoring of organotypic brain slices.

Brain slice preparations cultured in vitro have long been used as a simplified model for studying brain development, electrophysiology, neurodegeneration and neuroprotection. In this paper an open fluidic system developed for improved long term culturing of organotypic brain slices is presented. The positive effect of continuous flow of growth medium, and thus stability of the glucose concentration and waste removal, is simulated and compared to the effect of stagnant medium that is most often used in tissue culturing. Furthermore, placement of the tissue slices in the developed device was studied by numerical simulations in order to optimize the nutrient distribution. The device was tested by culturing transverse hippocampal slices from 7 days old NMRI mice for a duration of 14 days. The slices were inspected visually and the slices cultured in the fluidic system appeared to have preserved their structure better than the control slices cultured using the standard interface method.

Endoluminal isoproterenol reduces renal pelvic pressure during semirigid ureterorenoscopy: a porcine model.

OBJECTIVE: To investigate the effects on the pressure-flow relation of renal pelvic pressure during semirigid ureterorenoscopy and endoluminal perfusion of isoproterenol (ISO) 0.1 microg/mL, with emphasis on local effects and cardiovascular side-effects, as topically administered ISO effectively and dose-dependently causes relaxation of the upper urinary tract in pigs with no concomitant cardiovascular side-effects. MATERIALS AND METHODS: In anaesthetized female pigs (60 kg), 16 macroscopically normal upper urinary tract systems were subjected to ureterorenoscopy. Via a subcostal incision a 6-F catheter was placed in the renal pelvis for pressure measurements, and a semirigid ureteroscope (7.8 F) was inserted retrogradely in the renal pelvis, through which the pelvis was perfused. The blood pressure and heart rate were recorded. The increase in renal pelvic pressure was examined with increasing flow rates (0, 4, 8, 12, 16, 25 and 33 mL/min) with saline alone or saline + ISO 0.1 microg/mL. Perfusion was initiated on the left side, with randomization for adding ISO or not. Thereafter perfusion was done on the right side as a control in each pig. The surgeons were unaware of whether ISO was added or not. RESULTS: The mean (sd) baseline pelvic pressures in the saline and ISO group were 28 (7.1) and 25 (9.8) mmHg, respectively, with no significant difference (P = 0.079). Endoluminal perfusion with ISO significantly inhibited the pelvic pressure increase to perfusion at all perfusion rates. The pressure-flow relation was linear; the maximum relaxation (27%) was obtained at 4 mL/min, from 52 to 38 mmHg during saline alone and ISO 0.1 microg/mL perfusion, respectively. The mean blood pressure did not change significantly (P = 0.330). The mean (sd) heart rate in the saline and ISO group were 109 (4.5) and 97 (2.1) beats/min, respectively (P < 0.001), i.e. a markedly greater rate in the saline than in the ISO group. CONCLUSION: The pressure-flow relation during semirigid ureterorenoscopy was linear. ISO 0.1 microg/mL in saline significantly reduced the pressure-flow relation during semirigid ureterorenoscopy in this porcine model. ISO might be a potential additive to the irrigation fluid during upper urinary tract endoscopic procedures, minimizing pressure increases due to irrigation and manipulation.

Age-dependent plasticity of sex pheromone response in the moth, Agrotis ipsilon: combined effects of octopamine and juvenile hormone.

Male moths use sex pheromones to find their mating partners. In the moth, Agrotis ipsilon, the behavioral response and the neuron sensitivity within the primary olfactory centre, the antennal lobe (AL), to sex pheromone increase with age and juvenile hormone (JH) biosynthesis. By manipulating the JH level, we previously showed that JH controls this age-dependent neuronal plasticity, and that its effects are slow (within 2 days). We hypothesized that the hormonal effect might be indirect, and one neuromodulator candidate, which might serve as a mediator, is octopamine (OA). Here, we studied the effects of OA and an OA receptor antagonist, mianserin, on behavioral and AL neuron responses of mature and immature males during stimulation with sex pheromone. Our results indicate that, although OA injections enhanced the behavioral pheromone response in mature males, OA had no significant effect on behavior in immature males. However, mianserin injections decreased the behavioral response in mature males. AL neuron sensitivity increased after OA treatment in immature males, and decreased after mianserin treatment in mature males. Determination of OA levels in ALs of immature and mature males did not reveal any difference. To study the possible interactive effects of JH and OA, the behavioral pheromone response was analyzed in JH-deprived mature males injected with OA, and in immature males injected with fenoxycarb, a JH agonist, and mianserin. Results show that both JH and OA are necessary to elicit a behavioral response of A. ipsilon males to sex pheromone.

Dopamine release in organotypic cultures of foetal mouse mesencephalon: effects of depolarizing agents, pargyline, nomifensine, tetrodotoxin and calcium.

Organotypic mesencephalic cultures provide an attractive in vitro alternative to study development of the nigrostriatal system and pathophysiological mechanisms related to Parkinson's disease. However, dopamine (DA) release mechanisms have been poorly characterized in such cultures. We report here endogenous DA release (assessed by high-performance liquid chromatography) in organotypic cultures of foetal mouse (E12) midbrain following single or multiple challenges (1-h incubations) with high K(+) or veratridine in the presence or absence of pargyline, nomifensine, calcium and/or tetrodotoxin (TTX). Basal (i.e. spontaneous) DA release was only detected in the presence of pargyline and nomifensine (PN), and was highly dependent on calcium and sensitive to TTX. Basal DA release increased 2.4-fold between week 3 (1st DA release experiment) and week 4 in vitro (3rd DA release experiment), DA tissue levels increased 1.6-fold and DA release expressed as a percentage of total DA (medium + tissue contents) increased from 20% to 34% during this growth period in vitro. Co-treatments with high K(+) or veratridine did not cause major changes in percentages of DA release. Tyrosine hydroxylase activity was increased by high K(+), but not by the other drug treatments. The acute (single or multiple) treatments with depolarizing agents did not affect the survival of dopaminergic neurons, but chronic low-level veratridine treatments were toxic.

Renal nerves and nNOS: roles in natriuresis of acute isovolumetric sodium loading in conscious rats.

It was hypothesized that renal sympathetic nerve activity (RSNA) and neuronal nitric oxide synthase (nNOS) are involved in the acute inhibition of renin secretion and the natriuresis following slow NaCl loading (NaLoad) and that RSNA participates in the regulation of arterial blood pressure (MABP). This was tested by NaLoad after chronic renal denervation with and without inhibition of nNOS by S-methyl-thiocitrulline (SMTC). In addition, the acute effects of renal denervation on MABP and sodium balance were assessed. Rats were investigated in the conscious, catheterized state, in metabolic cages, and acutely during anesthesia. NaLoad was performed over 2 h by intravenous infusion of hypertonic solution (50 micromol.min(-1).kg body mass(-1)) at constant body volume conditions. SMTC was coinfused in amounts (20 microg.min(-1).kg(-1)) reported to selectively inhibit nNOS. Directly measured MABPs of acutely and chronically denervated rats were less than control (15% and 9%, respectively, P < 0.005). Plasma renin concentration (PRC) was reduced by renal denervation (14.5 +/- 0.2 vs. 19.3 +/- 1.3 mIU/l, P < 0.005) and by nNOS inhibition (12.4 +/- 2.3 vs. 19.6 +/- 1.6 mlU/l, P < 0.005). NaLoad reduced PRC (P < 0.05) and elevated MABP modestly (P < 0.05) and increased sodium excretion six-fold, irrespective of renal denervation and SMTC. The metabolic data demonstrated that renal denervation lowered sodium balance during the first days after denervation (P < 0.001). These data show that renal denervation decreases MABP and renin secretion. However, neither renal denervation nor nNOS inhibition affects either the renin down-regulation or the natriuretic response to acute sodium loading. Acute sodium-driven renin regulation seems independent of RSNA and nNOS under the present conditions.

Expansion and characterization of ventral mesencephalic precursor cells: effect of mitogens and investigation of FA1 as a potential dopaminergic marker.

Methods for identification and in vitro expansion of ventral mesencephalic dopaminergic precursor cells are of interest in the search for transplantable neurons for cell therapy in Parkinson's disease (PD). We investigated the potential use of fibroblast growth factor 2 (FGF2) and fibroblast growth factor 8 (FGF8) for expansion of such dopaminergic precursor cells, and fetal antigen-1 (FA1), a secreted neuronal protein of unknown function, as a non-invasive dopaminergic marker. Tissue from embryonic day (ED) 12 rat ventral mesencephalon was dissociated mechanically and cultured for 4 days in the presence of FGF2, FGF8, or without mitogens (control). After mitogen withdrawal and addition of 0.5% bovine serum, cells were differentiated for 6 days. Before differentiation, significantly more cells incorporated BrdU in cultures exposed to FGF2 (19-fold; P < 0.001) and FGF8 (3-fold; P < 0.05) compared to controls. After differentiation, biochemical analyses showed significantly more dopamine and FA1 in conditioned medium from both FGF2 and FGF8 expanded cultures than in controls. Correspondingly, numbers of tyrosine hydroxylase (TH)- and FA1-immunoreactive cells had increased 16-fold (P < 0.001) and 2.1-fold (P < 0.001), respectively in the FGF2 group and 10-fold (P < 0.001) and 1.8-fold (P < 0.05), respectively in the FGF8 group. In conclusion, the present procedure allows efficient expansion and differentiation of dopaminergic precursor cells and provides novel evidence of FGF8 as a mitogen for these cells. Furthermore, FA1 was identified as a potential supplementary non-invasive marker of cultured dopaminergic neurons.