The Smoothened agonist SAG Modulates the Male and Female Peripheral Immune Systems Differently in an Immune Model of Central Nervous System Demyelination.

Both Hedgehog and androgen signaling pathways are known to promote myelin regeneration in the central nervous system. Remarkably, the combined administration of agonists of each pathway revealed their functional cooperation towards higher regeneration in demyelination models in males. Since multiple sclerosis, the most common demyelinating disease, predominates in women, and androgen effects were reported to diverge according to sex, it seemed essential to assess the existence of such cooperation in females. Here, we developed an intranasal formulation containing the Hedgehog signaling agonist SAG, either alone or in combination with testosterone. We show that SAG promotes myelin regeneration and presumably a pro-regenerative phenotype of microglia, thus mimicking the effects previously observed in males. However, unlike in males, the combined molecules failed to cooperate in the demyelinated females, as shown by the level of functional improvement observed. Consistent with this observation, SAG administered in the absence of testosterone amplified peripheral inflammation by presumably activating NK cells and thus counteracting a testosterone-induced reduction in Th17 cells when the molecules were combined. Altogether, the data uncover a sex-dependent effect of the Hedgehog signaling agonist SAG on the peripheral innate immune system that conditions its ability to cooperate or not with androgens in the context of demyelination.

Testosterone Inhibits Secretion of the Pro-Inflammatory Chemokine CXCL1 from Astrocytes.

Astrocytes play an important role in the regulation of the inflammatory response in the CNS, e.g., in demyelinating diseases. Since the chemokine CXCL1 is known to be secreted by astrocytes and to have a pro-inflammatory effect on immune cells in the CNS, we verified the effect of testosterone on its secretion in vitro (in the astrocytic cell line DI TNC1). Testosterone reduced the increase in CXCL1 production caused by the pro-inflammatory agent lysophosphatidylcholine and restored the basal production level of CXCL1. The androgen receptor (present and functional in the studied cell line) was strongly suggested to mediate this effect-its non-steroid ligand flutamide exerted an agonist-like effect, mimicking the activity of testosterone itself on CXCL1 secretion. This novel mechanism has important implications for the known immunomodulatory effect of testosterone and potentially other androgenic hormones. It provides a potential explanation on the molecular level and shows that astrocytes are important players in inflammatory homeostasis in the CNS and its hormonal regulation. Therefore, it suggests new directions for the development of the therapeutic intervention.

Androgens show sex-dependent differences in myelination in immune and non-immune murine models of CNS demyelination.

Neuroprotective, anti-inflammatory, and remyelinating properties of androgens are well-characterized in demyelinated male mice and men suffering from multiple sclerosis. However, androgen effects mediated by the androgen receptor (AR), have been only poorly studied in females who make low androgen levels. Here, we show a predominant microglial AR expression in demyelinated lesions from female mice and women with multiple sclerosis, but virtually undetectable AR expression in lesions from male animals and men with multiple sclerosis. In female mice, androgens and estrogens act in a synergistic way while androgens drive microglia response towards regeneration. Transcriptomic comparisons of demyelinated mouse spinal cords indicate that, regardless of the sex, androgens up-regulate genes related to neuronal function integrity and myelin production. Depending on the sex, androgens down-regulate genes related to the immune system in females and lipid catabolism in males. Thus, androgens are required for proper myelin regeneration in females and therapeutic approaches of demyelinating diseases need to consider male-female differences.

Nose-to-Brain (N2B) Delivery: An Alternative Route for the Delivery of Biologics in the Management and Treatment of Central Nervous System Disorders.

In recent years, there have been a growing number of small and large molecules that could be used to treat diseases of the central nervous system (CNS). Nose-to-brain delivery can be a potential option for the direct transport of molecules from the nasal cavity to different brain areas. This review aims to provide a compilation of current approaches regarding drug delivery to the CNS via the nose, with a focus on biologics. The review also includes a discussion on the key benefits of nasal delivery as a promising alternative route for drug administration and the involved pathways or mechanisms. This article reviews how the application of various auxiliary agents, such as permeation enhancers, mucolytics, in situ gelling/mucoadhesive agents, enzyme inhibitors, and polymeric and lipid-based systems, can promote the delivery of large molecules in the CNS. The article also includes a discussion on the current state of intranasal formulation development and summarizes the biologics currently in clinical trials. It was noted that significant progress has been made in this field, and these are currently being applied to successfully transport large molecules to the CNS via the nose. However, a deep mechanistic understanding of this route, along with the intimate knowledge of various excipients and their interactions with the drug and nasal physiology, is still necessary to bring us one step closer to developing effective formulations for nasal-brain drug delivery.

Dissolution Improvement of Progesterone and Testosterone via Impregnation on Mesoporous Silica Using Supercritical Carbon Dioxide.

Progesterone (PRG) and testosterone (TST) were impregnated on mesoporous silica (ExP) particles via supercritical carbon dioxide (scCO2) processing at various pressures (10-18 MPa), temperatures (308.2-328.2 K), and time (30-360 min). The impact of a co-solvent on the impregnation was also studied at the best determined pressure and temperature. The properties of the drug embedded in silica particles were analysed via gas chromatography (GC), attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopy, X-ray diffraction (XRD), differential scanning calorimetry (DSC), and nitrogen adsorption. An impregnation of 1 to 82 mg/g for PRG and 0.1 to 16 mg/g for TST was obtained depending on the processing parameters. There was a significant effect of pressure, time, and co-solvent on the impregnation efficiency. Generally, an increase in time and pressure plus the use of co-solvent led to an improvement in drug adsorption. Conversely, a rise in temperature resulted in lower impregnation of both TST and PRG on ExP. There was a substantial increase in the dissolution rate (> 90% drug release within the first 2 min) of both TST and PRG impregnated in silica particles when compared to the unprocessed drugs. This dissolution enhancement was attributed to the amorphisation of both drugs due to their adsorption on mesoporous silica.

D2-like receptor activation by intranasal dopamine attenuates fear responses induced by electrical stimulation of the dorsal periaqueductal grey matter, but fails to reduce aversion to pit vipers and T-maze performance.

BACKGROUND: Panic-like reactions elicited by electrical stimulation of the dorsal periaqueductal grey matter (ES-dPAG) seem to be regulated by dopamine (DA). We showed that DA applied intranasally (IN) increased escape-behaviour thresholds induced by ES-dPAG of rats, indicating a panicolytic-like effect. AIMS: We investigated whether IN-DA increases escape-response thresholds induced by ES-dPAG by acting on D2-like receptors, and whether IN-DA affects escape responses elicited by the presence of a potential predator and by open space and height of the elevated T-maze (ETM) as well as motor performance in the open field (OF) test. METHODS: Wistar rats exposed to ES-dPAG were treated with Sulpiride (SUL, 40 mg/kg, D2-like receptor antagonist) previously IN-DA (2 mg/kg). Independent groups of rats treated with IN-DA were submitted to prey versus snake paradigm (PSP), ETM and OF. RESULTS: Anti-aversive effects of the IN-DA were reduced by SUL pretreatment in the ES-dPAG test. IN-DA did not affect the escape number in the PSP nor the escape latencies in the ETM as well as motor performance in the OF. CONCLUSIONS/INTERPRETATION: The IN-DA effects in reducing unconditioned fear responses elicited by ES-dPAG seem to be mediated by D2-like receptors. The lack of effects on panic-related responses in the ETM and PSP may be related to the possibility of avoiding the danger inherent to these models, a defence strategy not available during ES-dPAG. These findings cannot be attributed to motor performance. The decision-making responses to avoid dangerous situations can be orchestrated by supra-mesencephalic structures connected by non-dopaminergic inputs.

Mutation of Proteolipid Protein 1 Gene: From Severe Hypomyelinating Leukodystrophy to Inherited Spastic Paraplegia.

Pelizaeus-Merzbacher Disease (PMD) is an inherited leukodystrophy affecting the central nervous system (CNS)-a rare disorder that especially concerns males. Its estimated prevalence is 1.45-1.9 per 100,000 individuals in the general population. Patients affected by PMD exhibit a drastic reduction or absence of myelin sheaths in the white matter areas of the CNS. The Proteolipid Protein 1 (PLP1) gene encodes a transmembrane proteolipid protein. PLP1 is the major protein of myelin, and it plays a key role in the compaction, stabilization, and maintenance of myelin sheaths. Its function is predominant in oligodendrocyte development and axonal survival. Mutations in the PLP1 gene cause the development of a wide continuum spectrum of leukopathies from the most severe form of PMD for whom patients exhibit severe CNS hypomyelination to the relatively mild late-onset type 2 spastic paraplegia, leading to the concept of PLP1-related disorders. The genetic diversity and the biochemical complexity, along with other aspects of PMD, are discussed to reveal the obstacles that hinder the development of treatments. This review aims to provide a clinical and mechanistic overview of this spectrum of rare diseases.

The activation of D2-like receptors by intranasal dopamine facilitates the extinction of contextual fear and prevents conditioned fear-induced antinociception.

Fear extinction (FExt) is used to treat patients with posttraumatic stress disorder (PTSD). However, fear related to traumatic events can be persistent and return even after successful extinction. The neurochemical control of extinction seems to be performed by several neurotransmitters, including dopamine (DA), through D1 and D2 receptors. Recently, we showed that intranasally applied DA (IN-DA) facilitated the FExt, but the mechanisms by which it promoted this effect are still unknown. This study focused on investigating whether these effects are mediated by the action of DA on D2-like receptors since these receptors seem to be related to neurochemical and molecular changes underlying extinction. Also, we investigated whether IN-DA treatment would affect conditioned fear-induced antinociception (Fear-IA). Rats treated with IN-DA (1 mg/kg) twenty-five minutes after sulpiride (SUL; 40 mg/kg, i.p., D2-antagonist) were subjected to the extinction of contextual fear. IN-DA applied before the extinction session induced the FExt and prevented Fear-IA. These effects were impaired by pre-treatment with SUL, suggesting that the IN-DA effects are mediated by DA on D2-like receptors. SUL per se also facilitated the FExt but did not affect Fear-IA. These data suggest IN-DA as a promising pharmacological tool to supplement the psychotherapy of patients suffering from PTSD.

Sex differences in the cerebroprotection by Nestorone intranasal delivery following stroke in mice.

Our previous studies showed that intranasal delivery of progesterone offers a good bioavailability and neuroprotective efficacy after experimental stroke. We have also demonstrated that progesterone receptors (PR) are essential for cerebroprotection by endogenous progesterone and by progesterone treatment. The identification of PR as a potential drug target for stroke therapy opens new therapeutic indications for selective synthetic progestins. Nestorone® (16-methylene-17α-acetoxy-19-nor-pregn-4-ene-3, 20-dione, also known as segesterone acetate) is a 19-norprogesterone derivative that more potently targets PR than progesterone. The objective of this study was to evaluate the cerebroprotective efficiency of intranasal administration of Nestorone after middle cerebral occlusion (MCAO) in mice. We show here that intranasal administration is a very efficient route to achieve a preferential delivery of Nestorone to the brain and confers a slow elimination and a sustained bioavailability. Furthermore, intranasal administration of Nestorone (at 0.08 mg/kg) improved the functional outcomes and decreased the ischemic lesion in male but not in female mice at 48 h post MCAO. Use of PRNesCre mice, selectively lacking expression of PR in neural cells, and their control PRloxP/loxP littermates showed that the cerebroprotective effects of Nestorone in male mice depended on neural PR as they were not observed in PRNesCre mice. Our findings show that intranasal delivery of Nestorone may be an efficient strategy to promote recovery after stroke in males and confirm the key role of PR in cerebroprotection. Furthermore, they point to sex differences in the response to Nestorone treatment and emphasize the necessity to include males and females in experimental studies.

Acute intranasal dopamine application counteracts the reversal learning deficit of spontaneously hypertensive rats in an attentional set-shifting task.

RATIONALE: Studies on the attention-deficit/hyperactivity disorder (ADHD) have concluded that the disorder might be caused by a deficit in the inhibitory control of executive functions because of dopamine hypofunction. Recently, the intranasal route has emerged as an effective alternative means for sending dopamine directly to the brain. However, whether the treatment can ameliorate the deficits of inhibitory control in ADHD remains unknown. OBJECTIVES: Investigating the effects of acute intranasal dopamine (IN-DA) on the inhibitory control of executive functions of an ADHD rodent model. METHODS: We trained an animal model of ADHD, the spontaneously hypertensive rat (SHR), and Wistar rats as controls, in an attentional set-shifting task (ASST) in which dopamine (0.15 mg/kg, 0.3 mg/kg, or vehicle) was intranasally administered before the final test. RESULTS: IN-DA application dose-dependently improved the performance and reduced errors of SHR in the initial reversal learning. The effect size was comparable to that of a peripheral injection of 0.6 mg/kg methylphenidate. In control Wistar rats, the highest dose of intranasal dopamine (0.3 mg/kg) induced deficits in the reversal learning of extradimensional discriminations. CONCLUSIONS: The findings suggest that the IN-DA treatment has potential for use in the treatment of ADHD; however, caution must be exercised when determining the dosage to be administered, because too much dopamine may have negative effects.

Functional cooperation of the hedgehog and androgen signaling pathways during developmental and repairing myelination.

Hedgehog morphogens control fundamental cellular processes during tissue development and regeneration. In the central nervous system (CNS), Hedgehog signaling has been implicated in oligodendrocyte and myelin production, where it functions in a concerted manner with other pathways. Since androgen receptor (AR) plays a key role in establishing the sexual phenotype of myelin during development and is required for spontaneous myelin regeneration in the adult CNS, we hypothesized the existence of a possible coordination between Hedgehog and androgen signals in oligodendrocyte and myelin production. Here, we report complementary activities of both pathways during early postnatal oligodendrogenesis further revealing that persistent Hedgehog signaling activation impedes myelin production. The data also uncover prominent pro-myelinating activity of testosterone and involvement of AR in the control of neural stem cell commitment toward the oligodendroglial lineage. In the context of CNS demyelination, we provide evidence for the functional cooperation of the pathways leading to acceleration of myelin regeneration that might be related to their respective role on microglial and astroglial responses, higher preservation of axonal integrity, lower neuroinflammation, and functional improvement of animals in an immune model of CNS demyelination. Strong decreases of deleterious cytokines in the CNS (GM-CSF, TNF-α, IL-17A) and spleen (IL-2, IFN-γ) stand as unique features of the combined drugs while the potent therapeutic activity of testosterone on peripheral immune cells contributes to increase tolerogenic CD11c+ dendritic cells, reduce the clonal expansion of conventional CD4+ T cells and increase CD4+ Foxp3+ regulatory T cells. Altogether, these data might open promising perspectives for demyelinating diseases.

Altered dopaminergic pathways and therapeutic effects of intranasal dopamine in two distinct mouse models of autism.

The dopamine (DA) system has a profound impact on reward-motivated behavior and is critically involved in neurodevelopmental disorders, such as autism spectrum disorder (ASD). Although DA defects are found in autistic patients, it is not well defined how the DA pathways are altered in ASD and whether DA can be utilized as a potential therapeutic agent for ASD. To this end, we employed a phenotypic and a genetic ASD model, i.e., Black and Tan BRachyury T+Itpr3tf/J (BTBR) mice and Fragile X Mental Retardation 1 knockout (Fmr1-KO) mice, respectively. Immunostaining of tyrosine hydroxylase (TH) to mark dopaminergic neurons revealed an overall reduction in the TH expression in the substantia nigra, ventral tegmental area and dorsal striatum of BTBR mice, as compared to C57BL/6 J wild-type ones. In contrast, Fmr1-KO animals did not show such an alteration but displayed abnormal morphology of TH-positive axons in the striatum with higher "complexity" and lower "texture". Both strains exhibited decreased expression of striatal dopamine transporter (DAT) and increased spatial coupling between vesicular glutamate transporter 1 (VGLUT1, a label for glutamatergic terminals) and TH signals, while GABAergic neurons quantified by glutamic acid decarboxylase 67 (GAD67) remained intact. Intranasal administration of DA rescued the deficits in non-selective attention, object-based attention and social approaching of BTBR mice, likely by enhancing the level of TH in the striatum. Application of intranasal DA to Fmr1-KO animals alleviated their impairment of social novelty, in association with reduced striatal TH protein. These results suggest that although the DA system is modified differently in the two ASD models, intranasal treatment with DA effectively rectifies their behavioral phenotypes, which may present a promising therapy for diverse types of ASD.

The Effect of a Thixotropic Nasal Gel on Nasal Symptoms and Inflammatory Biomarkers in Seasonal Allergic Rhinitis.

BACKGROUND: Drug-free viscous nasal applications have been shown to reduce nasal symptoms in individuals with seasonal allergic rhinitis (SAR). Nascum®-Plus (NP), a commercially available thixotropic gel, has been designed to reduce dryness and soreness of the nasal mucosa and prevent the absorption of small particles. OBJECTIVES: The aim of this study was to assess the efficacy of single-dose NP in treating nasal symptoms and secretion during challenge in an allergen challenge chamber (ACC). Furthermore, the effect of this treatment on biomarkers and immune cells of the allergic cascade were measured. METHODS: This open-label, cross-over, sequence-randomized, monocentric trial randomized 18 adults with SAR and a positive skin prick test reaction to Dactylis glomerata pollen to receive NP or no treatment during two 4-h ACC sessions 3 weeks apart. On Day 1, 9 subjects were challenged for 4 h with treatment, the other 9 without treatment, and vice versa on Day 22. Nasal lavage fluid and nasal filter eluate samples were obtained pre, 2, and 18 h post challenge in the ACC. RESULTS: NP significantly reduced nasal symptoms, assessed by total nasal symptom score (p < 0.001), and minimized nasal secretion (p = 0.047), while no significant effect on biomarkers and immune cells in the nasal fluid was observed. The treatment was safe and well-tolerated. CONCLUSIONS: The physical barrier built by NP nasal gel can be safely applied in patients with allergic rhinitis. It reduces allergic nasal symptoms and secretion, but application of a single dose does not affect local inflammatory biomarkers.

Dose-dependent and long-term cerebroprotective effects of intranasal delivery of progesterone after ischemic stroke in male mice.

Intranasal administration is emerging as a very promising route to deliver therapeutics to the brain. We have recently shown that the intranasal delivery of progesterone at 8 mg/kg is neuroprotective after stroke in male mice. To explore the translational potential of intranasal progesterone treatment, we performed a dose-response study and analyzed outcomes at 48 h after middle cerebral artery occlusion (MCAO). The effects on functional outcomes at long-term were examined by using the optimal dose. In the first experiment, male C57BL/6JRj mice were treated with progesterone at 8, 16 or 24 mg/kg, or with placebo at 1, 6 and 24 h post-MCAO. Our results show that the dose of 8 mg/kg was optimal in counteracting the early histopathological impairments as well as in improving functional recovery. Steroid profiling in plasma showed that the dose of 8 mg/kg is the one that leads to sustained high levels of progesterone and its neuroactive metabolites. In the second experiment, the dose of 8 mg/kg was used and analyzes were performed at 2, 7 and 21 days post-MCAO. Progesterone increased survival, glycemia and body weight. Furthermore, progesterone decreased neurological deficits and improved performances of mice on the rotarod and pole as early as 2 days and up to 21 days post-MCAO. These findings show that intranasal administration of progesterone has a significant translational potential as a cerebroprotective treatment after stroke that can be effective to reduce mortality, to limit tissue and cell damage at the acute phase; and to confer a long-term functional recovery.

Intranasal pregnenolone increases acetylcholine in frontal cortex, hippocampus, and amygdala-Preferentially in the hemisphere ipsilateral to the injected nostril.

This study determined the effects of intranasal pregnenolone (IN-PREG) on acetylcholine (ACh) levels in selected areas of the rat brain, using in vivo microdialysis. Previous studies showed that PREG rapidly reaches the rodent brain after intranasal administration and that direct infusion of PREG and PREG-S into the basal forebrain modulates ACh release in frontal cortex, amygdala, and hippocampus. In the present study, we investigated the effects of IN-PREG on the cholinergic system in the rat brain. In the first experiment, IN-PREG (5.6 and 11.2 mg/ml) or vehicle was applied bilaterally, and we hypothesized that IN-PREG would increase ACh levels in amygdala, hippocampus, and frontal cortex, relative to baseline and vehicle. Dialysate was collected for 100 min, based on pilot data of duration of effect. Bilateral IN-PREG (5.6 and 11.2 mg/ml) increased frontal cortex and hippocampal ACh relative to both baseline and vehicle. Moreover, 11.2 mg/ml PREG increased ACh in the amygdala relative to baseline, the lower dose, and vehicle. Therefore, in the second experiment, IN-PREG (11.2 mg/ml) was applied only into one nostril, with vehicle applied into the other nostril, in order to determine whether ACh is predominantly increased in the ipsilateral relative to the contralateral amygdala. Unilateral application of IN-PREG increased ACh in the ipsilateral amygdala, whereas no effect was observed on the contralateral side, suggesting that PREG was transported from the nostrils to the brain via the olfactory epithelial pathway, but not by circulation. The present data provide additional information on IN-PREG action in the cholinergic system of frontal cortex, amygdala, and hippocampus. This may be relevant for therapeutic IN application of PREG in neurogenerative and neuropsychiatric disorders.

Intranasal dopamine attenuates fear responses induced by electric shock to the foot and by electrical stimulation of the dorsal periaqueductal gray matter.

PURPOSE: Intranasally applied dopamine (IN-DA), which likely reaches the brain via nasal-brain pathways and bypasses the blood-brain barrier, has been found to increase extracellular DA and bind to the DA2 transporter in the striatum. Recent studies suggest that DA plays a significant role in the processing of signaled and unconditioned aversive stimulation, including evidence that may attenuate responses to painful input. The purpose of this study was to examine the effects of IN-DA on fear-related behaviors induced by electric shock to the foot or by electrical stimulation of the dorsal periaqueductal gray matter (dPAG). METHODS: DA hydrochloride suspended in a viscous castor oil gel (1 or 2 mg/kg) was applied (IN-DA) in a volume of 5 µL into the nostrils of adult Wistar male rats in order to evaluate its effects on (a) freezing induced by electric shock to the foot and (b) thresholds of freezing and escape and duration of post-stimulation freezing induced by electrical stimulation of the dPAG. RESULTS: IN-DA attenuated freezing induced by electric shock to the foot in the three test trials, indicating that it reduced long-term fear responses. IN-DA also increased the threshold of dPAG stimulation-induced escape responses and reduced post-stimulation freezing. CONCLUSIONS: IN-DA, which has previously been shown to facilitate learning and to have antidepressive-like effects, attenuated unconditioned fear responses elicited by peripheral and intramesencephalic (dPAG) stimulation and reduced long-term conditioned fear responses.

Intranasal administration of progesterone: A potential efficient route of delivery for cerebroprotection after acute brain injuries.

Progesterone has been shown to be cerebroprotective in different experimental models of brain injuries and neurodegenerative diseases. The preclinical data provided great hope for its use in humans. The failure of Phase 3 clinical trials to demonstrate the cerebroprotective efficiency of progesterone in traumatic brain injury (TBI) patients emphasizes that different aspects of the design of both experimental and clinical studies should be reviewed and refined. One important aspect to consider is to test different routes of delivery of therapeutic agents. Several studies have shown that the intranasal delivery of drugs could be used in different experimental models of central nervous system diseases. In this review, we will summarize the pharmacokinetic characteristics and practical advantages of intranasal delivery of progesterone. A special emphasis will be placed on describing and discussing our recent findings showing that intranasal delivery of progesterone after transient focal cerebral ischemia: 1) improved motor functions; 2) reduced infarct volume, neuronal loss, blood brain barrier disruption; and 3) reduced brain mitochondrial dysfunctions. Our data suggest that intranasal delivery of progesterone is a potential efficient, safe and non-stressful mode of administration that warrants evaluation for cerebroprotection in patients with brain injuries. This article is part of the Special Issue entitled "Novel Treatments for Traumatic Brain Injury".

Intra-nasal dopamine alleviates cognitive deficits in tgDISC1 rats which overexpress the human DISC1 gene.

The Disrupted-in-Schizophrenia 1 (DISC1) gene has been associated with mental illnesses such as major depression and schizophrenia. The transgenic DISC1 (tgDISC1) rat, which overexpresses the human DISC1 gene, is known to exhibit deficient dopamine (DA) homeostasis. To ascertain whether the DISC1 gene also impacts cognitive functions, 14-15 months old male tgDISC1 rats and wild-type controls were subjected to the novel object preference (NOP) test and the object-based attention test (OBAT) in order to assess short-term memory (1 h), long-term memory (24 h), and attention. RESULTS: The tgDISC1 group exhibited intact short-term memory, but deficient long-term-memory in the NOP test and deficient attention-related behavior in the OBAT. In a different group of tgDISC1 rats, 3 mg/kg intranasally applied dopamine (IN-DA) or its vehicle was applied prior to the NOP or the OBAT test. IN-DA reversed cognitive deficits in both the NOP and OBAT tests. In a further cohort of tgDISC1 rats, post-mortem levels of DA, noradrenaline, serotonin and acetylcholine were determined in a variety of brain regions. The tgDISC1 group had less DA in the neostriatum, hippocampus and amygdala, less acetylcholine in neostriatum, nucleus accumbens, hippocampus, and amygdala, more serotonin in the nucleus accumbens, and less serotonin and noradrenaline in the amygdala. CONCLUSIONS: Our findings show that DISC1 overexpression and misassembly is associated with deficits in long-term memory and attention-related behavior. Since behavioral impairments in tgDISC1 rats were reversed by IN-DA, DA deficiency may be a major cause for the behavioral deficits expressed in this model.

A Role of Endogenous Progesterone in Stroke Cerebroprotection Revealed by the Neural-Specific Deletion of Its Intracellular Receptors.

Treatment with progesterone protects the male and female brain against damage after middle cerebral artery occlusion (MCAO). However, in both sexes, the brain contains significant amounts of endogenous progesterone. It is not known whether endogenously produced progesterone enhances the resistance of the brain to ischemic insult. Here, we used steroid profiling by gas chromatography-tandem mass spectrometry (GC-MS/MS) for exploring adaptive and sex-specific changes in brain levels of progesterone and its metabolites after MCAO. We show that, in the male mouse brain, progesterone is mainly metabolized via 5α-reduction leading to 5α-dihydroprogesterone (5α-DHP), also a progesterone receptor (PR) agonist ligand in neural cells, then to 3α,5α-tetrahydroprogesterone (3α,5α-THP). In the female mouse brain, levels of 5α-DHP and 3α,5α-THP are lower and levels of 20α-DHP are higher than in males. After MCAO, levels of progesterone and 5α-DHP are upregulated rapidly to pregnancy-like levels in the male but not in the female brain. To assess whether endogenous progesterone and 5α-DHP contribute to the resistance of neural cells to ischemic damage, we inactivated PR selectively in the CNS. Deletion of PR in the brain reduced its resistance to MCAO, resulting in increased infarct volumes and neurological deficits in both sexes. Importantly, endogenous PR ligands continue to protect the brain of aging mice. These results uncover the unexpected importance of endogenous progesterone and its metabolites in cerebroprotection. They also reveal that the female reproductive hormone progesterone is an endogenous cerebroprotective neurosteroid in both sexes.SIGNIFICANCE STATEMENT The brain responds to injury with protective signaling and has a remarkable capacity to protect itself. We show here that, in response to ischemic stroke, levels of progesterone and its neuroactive metabolite 5α-dihydroprogesterone are upregulated rapidly in the male mouse brain but not in the female brain. An important role of endogenous progesterone in cerebroprotection was demonstrated by the conditional inactivation of its receptor in neural cells. These results show the importance of endogenous progesterone, its metabolites, and neural progesterone receptors in acute cerebroprotection after stroke. This new concept could be exploited therapeutically by taking into account the progesterone status of patients and by supplementing and reinforcing endogenous progesterone signaling for attaining its full cerebroprotective potential.

Intranasal administration of dopamine attenuates unconditioned fear in that it reduces restraint-induced ultrasound vocalizations and escape from bright light.

BACKGROUND: Although substantial evidence suggests that dopamine (DA) enhances conditioned fear responses, few studies have examined the role of DA in unconditioned fear states. Whereas DA does not cross the blood-brain barrier, intranasally-applied dopamine reaches the brain directly via the nose-brain pathways in rodents, providing an alternative means of targeting DA receptors. Intranasal dopamine (IN-DA) has been demonstrated to bind to DA transporters and to increase extracellular DA in the striatum as well as having memory-promoting effects in rats. The purpose of this study was to examine the influence of IN-DA in three tests of fear/anxiety. METHODS: The three doses of DA hydrochloride (0.03, 0.3, or 1 mg/kg) were applied in a viscous castor oil gel in a volume of 5 µl to each of both nostrils of adult Wistar rats prior to testing of (a) escape from a bright light, using a two-chamber procedure, (b) restraint-induced 22 kHz ultrasound vocalizations (USVs), and (c) exploratory behavior in the elevated plus-maze (EPM). RESULTS: IN-DA dose-dependently reduced escape from bright light and the number of USV responses to restraint. It had no influence on the exploratory behavior in the EPM. CONCLUSIONS: IN-DA application reduced escape behavior in two tests of unconditioned fear (escape from bright light and USV response to immobilization). These findings may be interpreted in light of the known antidepressant action of IN-DA and DA reuptake blockers. The results also confirm the promise of the nasal route as an alternative means for targeting the brain's dopaminergic receptors with DA.