Reduced light exposure mitigates streptozotocin-induced vascular changes and gliosis in diabetic retina by an anti-inflammatory effect and increased retinal cholesterol turnover.

Diabetic retinopathy is not cured efficiently and changes of lifestyle measures may delay early retinal injury in diabetes. The aim of our study was to investigate the effects of reduced daily light exposure on retinal vascular changes in streptozotocin (STZ)-induced model of DM with emphasis on inflammation, Aqp4 expression, visual cycle and cholesterol metabolism-related gene expression in rat retina and RPE. Male Wistar rats were divided into the following groups: 1. control; 2. diabetic group (DM) treated with streptozotocin (100 mg/kg); 3. group exposed to light/dark cycle 6/18 h (6/18); 4. diabetic group exposed to light/dark cycle 6/18 h (DM+6/18). Retinal vascular abnormalities were estimated based on lectin staining, while the expression of genes involved in the visual cycle, cholesterol metabolism, and inflammation was determined by qRT-PCR. Reduced light exposure alleviated vasculopathy, gliosis and the expression of IL-1 and TNF-α in the retina with increased perivascular Aqp4 expression. The expression of genes involved in visual cycle and cholesterol metabolism was significantly up-regulated in RPE in DM+6/18 vs. DM group. In the retina only the expression of APOE was significantly higher in DM+6/18 vs. DM group. Reduced light exposure mitigates vascular changes and gliosis in DM via its anti-inflammatory effect, increased retinal cholesterol turnover and perivascular Aqp4 expression.

Nanocarriers in topical photodynamic therapy.

INTRODUCTION: Photodynamic therapy (PDT) has gained significant attention due to its superiority over conventional treatments. In the context of skin cancers and nonmalignant skin diseases, topical application of photosensitizer formulations onto affected skin, followed by illumination, offers distinct advantages. Topical PDT simplifies therapy by providing easy access to the skin, increasing drug concentration within the target area, and confining residual photosensitivity to the treated skin. However, the effectiveness of topical PDT is often hindered by challenges such as limited skin penetration or photosensitizer instability. Additionally, the hypoxic tumor environment poses further limitations. Nanocarriers present a promising solution to address these challenges. AREAS COVERED: The objective of this review is to comprehensively explore and highlight the role of various nanocarriers in advancing topical PDT for the treatment of skin diseases. The primary focus is to address the challenges associated with conventional topical PDT approaches and demonstrate how nanotechnology-based strategies can overcome these challenges, thereby improving the overall efficiency and efficacy of PDT. EXPERT OPINION: Nanotechnology has revolutionized the field of PDT, offering innovative tools to combat the unfavorable features of photosensitizers and hurdles in PDT. Nanocarriers enhance skin penetration and stability of photosensitizers, provide controlled drug release, reduce needed dose, increase production of reactive oxygen species, while reducing side effects, thereby improving PDT effectiveness.

The presymptomatic treatment with 3HFWC nanosubstance decreased plaque load in 5XFAD mouse model of Alzheimer's disease.

INTRODUCTION: In the present study, we assessed the effects of the hyper-harmonized-hydroxylated fullerene-water complex (3HFWC) on Alzheimer's disease (AD) neuropathological hallmarks in 5XFAD mice, an AD animal model. METHODS: The 3-week-old 5XFAD mice were exposed to 3HFWC water solution ad libitum for 3 months in the presymptomatic phase of pathology. The functional effects of the treatment were confirmed through near-infrared spectroscopy (NIRS) analysis through machine learning (ML) using artificial neural networks (ANNs) to classify the control and 3HFWC-treated brain tissue samples. The effects of 3HFWC treatment on amyloid-ß (Aß) accumulation, plaque formation, gliosis, and synaptic plasticity in cortical and hippocampal tissue were assessed. RESULTS: The 3HFWC treatment significantly decreased the amyloid-ß plaque load in specific parts of the cerebral cortex. At the same time, 3HFWC treatment did not induce the activation of glia (astrocytes and microglia) nor did it negatively affect synaptic protein markers (GAP-43, synaptophysin, and PSD-95). CONCLUSION: The obtained results point to the potential of 3HFWC, when applied in the presymptomatic phase of AD, to interfere with amyloid plaque formation without inducing AD-related pathological processes such as neuroinflammation, gliosis, and synaptic vulnerability.

Prolonged Zaleplon Treatment Increases the Expression of Proteins Involved in GABAergic and Glutamatergic Signaling in the Rat Hippocampus.

Zaleplon is a positive allosteric modulator of the γ-aminobutyric acid (GABA)A receptor approved for the short-term treatment of insomnia. Previous publications on zaleplon have not addressed the proteins involved in its mechanism of action but have mostly referred to behavioral or pharmacological studies. Since both GABAergic and glutamatergic signaling have been shown to regulate wakefulness and sleep, we examined the effects of prolonged zaleplon treatment (0.625 mg/kg for 5 days) on these systems in the hippocampus of male Wistar rats. Western blot and immunohistochemical analyses showed that the upregulated components of GABAergic signaling (glutamate decarboxylase, vesicular GABA transporter, GABA, and α1 subunit of the GABAA receptor) were accompanied by increased protein levels in the glutamatergic system (vesicular glutamate transporter 1 and NR1, NR2A, and NR2B subunits of N-methyl-d-aspartate receptor). Our results, showing that zaleplon enhances GABA neurotransmission in the hippocampus, were not surprising. However, we found that treatment also increased glutamatergic signaling. This could be the result of the downregulation of adenosine A1 receptors, important modulators of the glutamatergic system. Further studies are needed to investigate the effects of the zaleplon-induced increase in hippocampal glutamatergic neurotransmission and the possible involvement of the adenosine system in zaleplon's mechanism of action.

The Expression of Major Facilitator Superfamily Domain-Containing Protein2a (Mfsd2a) and Aquaporin 4 Is Altered in the Retinas of a 5xFAD Mouse Model of Alzheimer's Disease.

Cerebral amyloid angiopathy (CAA) is characterized by amyloid ß (Aß) accumulation in the blood vessels and is associated with cognitive impairment in Alzheimer's disease (AD). The increased accumulation of Aß is also present in the retinal blood vessels and a significant correlation between retinal and brain amyloid deposition was demonstrated in living patients and animal AD models. The Aß accumulation in the retinal blood vessels can be the result of impaired transcytosis and/or the dysfunctional ocular glymphatic system in AD and during aging. We analyzed the changes in the mRNA and protein expression of major facilitator superfamily domain-containing protein2a (Mfsd2a), the major regulator of transcytosis, and of Aquaporin4 (Aqp4), the key player implicated in the functioning of the glymphatic system, in the retinas of 4- and 12-month-old WT and 5xFAD female mice. A strong decrease in the Mfsd2a mRNA and protein expression was observed in the 4 M and 12 M 5xFAD and 12 M WT retinas. The increase in the expression of srebp1-c could be at least partially responsible for the Mfsd2a decrease in the 4 M 5xFAD retinas. The decrease in the pericyte (CD13+) coverage of retinal blood vessels in the 4 M and 12 M 5xFAD retinas and in the 12 M WT retinas suggests that pericyte loss could be associated with the Mfsd2a downregulation in these experimental groups. The observed increase in Aqp4 expression in 4 M and 12 M 5xFAD and 12 M WT retinas accompanied by the decreased perivascular Aqp4 expression is indicative of the impaired glymphatic system. The findings in this study reveal the impaired Mfsd2a and Aqp4 expression and Aqp4 perivascular mislocalization in retinal blood vessels during physiological (WT) and pathological (5xFAD) aging, indicating their importance as putative targets for the development of new treatments that can improve the regulation of transcytosis or the function of the glymphatic system.

Shortened Daily Photoperiod Alleviates Anxiety-like Behaviour by Antioxidant Effect and Changes Serum Fatty Acid Profile in Diabetic Rats.

The aim of our study was to investigate the effects of a shortened daily photoperiod on anxiety-like behaviour, brain oxidative stress, lipid status and fatty acid composition of serum lipids in a streptozotocin (STZ)-induced model of diabetes mellitus in rats. Male Wistar rats were divided into the following groups: first group-control group (C12/12); second group-diabetic group (DM12/12; 100 mg/kg STZ); third group-control group exposed to a light/dark cycle 6/18 h (C6/18); fourth group-diabetic group exposed to a light/dark cycle 6/18 h (DM6/18). Anxiety-like behaviour was tested three weeks following STZ injection by elevated plus maze (EPM) and open-field test (OFT). Oxidative stress parameters were measured in the cortex, hippocampus and thalamus, while lipid status and fatty acid methyl esters (FAMEs) were measured in the serum. Both EPM and OFT showed a lower degree of anxiety-like behaviour in the DM6/18 vs. DM12/12 group. Lipid peroxidation in the cortex, hippocampus and thalamus was significantly lower in the DM6/18 vs. DM12/12 group (p < 0.05), associated with an increased level of antioxidant enzymes and protein thiols in the cortex and thalamus. In the DM6/18 group, oleic, vaccenic, dihomo-γ-linolenic and docosahexaenoic acid concentrations were significantly higher in comparison to the DM12/12 group. A shortened daily photoperiod alleviates anxiety-like behaviour in diabetic rats by reduced lipid peroxidation and changes in the serum fatty acids profile.

Contribution of the opioid system to depression and to the therapeutic effects of classical antidepressants and ketamine.

Major depressive disorder (MDD) afflicts approximately 5 % of the world population, and about 30-50 % of patients who receive classical antidepressant medications do not achieve complete remission (treatment resistant depressive patients). Emerging evidence suggests that targeting opioid receptors mu (MOP), kappa (KOP), delta (DOP), and the nociceptin/orphanin FQ receptor (NOP) may yield effective therapeutics for stress-related psychiatric disorders. As depression and pain exhibit significant overlap in their clinical manifestations and molecular mechanisms involved, it is not a surprise that opioids, historically used to alleviate pain, emerged as promising and effective therapeutic options in the treatment of depression. The opioid signaling is dysregulated in depression and numerous preclinical studies and clinical trials strongly suggest that opioid modulation can serve as either an adjuvant or even an alternative to classical monoaminergic antidepressants. Importantly, some classical antidepressants require the opioid receptor modulation to exert their antidepressant effects. Finally, ketamine, a well-known anesthetic whose extremely efficient antidepressant effects were recently discovered, was shown to mediate its antidepressant effects via the endogenous opioid system. Thus, although opioid system modulation is a promising therapeutical venue in the treatment of depression further research is warranted to fully understand the benefits and weaknesses of such approach.

The supplementation of a high dose of fish oil during pregnancy and lactation led to an elevation in Mfsd2a expression without any changes in docosahexaenoic acid levels in the retina of healthy 2-month-old mouse offspring.

Introduction: During fetal development, the proper development of neural and visual systems relies on the maternal supplementation of omega-3 fatty acids through placental transfer. Pregnant women are strongly advised to augment their diet with additional sources of omega-3, such as fish oil (FO). This supplementation has been linked to a reduced risk of preterm birth, pre-eclampsia, and perinatal depression. Recently, higher doses of omega-3 supplementation have been recommended for pregnant women. Considering that omega-3 fatty acids, particularly docosahexaenoic acid (DHA), play a crucial role in maintaining the delicate homeostasis required for the proper functioning of the retina and photoreceptors the effects of high-dose fish oil (FO) supplementation during pregnancy and lactation on the retina and retinal pigmented epithelium (RPE) in healthy offspring warrant better understanding. Methods: The fatty acid content and the changes in the expression of the genes regulating cholesterol homeostasis and DHA transport in the retina and RPE were evaluated following the high-dose FO supplementation. Results: Our study demonstrated that despite the high-dose FO treatment during pregnancy and lactation, the rigorous DHA homeostasis in the retina and RPE of the two-month-old offspring remained balanced. Another significant finding of this study is the increase in the expression levels of major facilitator superfamily domain-containing protein (Mfsd2a), a primary DHA transporter. Mfsd2a also serves as a major regulator of transcytosis during development, and a reduction in Mfsd2a levels poses a major risk for the development of leaky blood vessels. Conclusion: Impairment of the blood-retinal barrier (BRB) is associated with the development of numerous ocular diseases, and a better understanding of how to manipulate transcytosis in the BRB during development can enhance drug delivery through the BRB or contribute to the repair of central nervous system (CNS) barriers.

Food Restriction Counteracts Dexamethasone-Induced Downregulation of Genes Involved in Cholesterol Homeostasis in Rat Brain during Aging.

Glucocorticoids are the most potent anti-inflammatory agents known. Limited in vivo data are available to characterize the mechanism underlying their cognitive side effects and transient occurrence of steroid psychosis. Cholesterol is important for proper neurotransmission and brain plasticity, and disruption of its homeostasis in the brain has been closely associated with memory decline during aging and in age-related neurodegenerative disorders. In the present study, we assessed the direct effects of dexamethasone, a potent synthetic glucocorticoid, on the expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), apolipoprotein E (ApoE) and cholesterol 24S-hydroxylase (CYP46A1), major enzymes involved in cholesterol synthesis, metabolism, and excretion, respectively. The effects of the dexamethasone were examined during aging, in the cortex and hippocampus of 6-, 12- and 18-month-old rats, and following long-term food restriction (FR). The most prominent change observed was the age-related decrease in ApoE mRNA regardless of the food regimen applied. In animals kept on FR, this decrease was accompanied by an increase in the mRNA expression of HMGCR and CYP46A1. The present study also demonstrates that food restriction reversed most of the dexamethasone-induced changes in the expression of genes involved in regulation of cholesterol homeostasis in aging rats, in a region-specific manner.

Tryptophan metabolites in depression: Modulation by gut microbiota.

Clinical depression is a multifactorial disorder and one of the leading causes of disability worldwide. The alterations in tryptophan metabolism such as changes in the levels of serotonin, kynurenine, and kynurenine acid have been implicated in the etiology of depression for more than 50 years. In recent years, accumulated evidence has revealed that gut microbial communities, besides being essential players in various aspects of host physiology and brain functioning are also implicated in the etiology of depression, particularly through modulation of tryptophan metabolism. Therefore, the aim of this review is to summarize the evidence of the role of gut bacteria in disturbed tryptophan metabolism in depression. We summed up the effects of microbiota on serotonin, kynurenine, and indole pathway of tryptophan conversion relevant for understanding the pathogenesis of depressive behavior. Moreover, we reviewed data regarding the therapeutic effects of probiotics, particularly through the regulation of tryptophan metabolites. Taken together, these findings can open new possibilities for further improvement of treatments for depression based on the microbiota-mediated modulation of the tryptophan pathway.

Fatty acids as biomodulators of Piezo1 mediated glial mechanosensitivity in Alzheimer's disease.

The brain is the softest organ in the body, and any change in the mechanical properties of the tissue induces the activation of glial cells, astrocytes and microglia. Amyloid plaques, one of the main pathological features of Alzheimer's disease (AD), are substantially harder than the surrounding brain tissue and can activate astrocytes and microglia resulting in the glial engulfment of plaques. Durotaxis, a migratory preference towards stiffer tissue, is prompting microglia to form a mechanical barrier around plaques reducing amyloid ß (Aß) induced neurotoxicity. Mechanoreceptors are highly expressed in the brain, particularly in microglia. The large increase in the expression of the mechanoreceptor Piezo1 was observed in the brains from AD animal models and AD patients in plaque encompassing glia. Importantly, Piezo1 function is regulated via force-from-lipids through the lipid composition of the membrane and membranous incorporation of polyunsaturated fatty acids (PUFAs) can affect the function of Piezo1 altering mechanosensitive properties of the cell. On the other hand, PUFAs dietary supplementation can alter microglial polarization, the envelopment of amyloid plaques, and immune response and Piezo1 activity was implicated in the similar modulations of microglia behavior. Finally, PUFAs treatment is currently in use in medical trials as the therapy for sickle cell anemia, a disease linked with the mutations in Piezo1. Further studies are needed to elucidate the connection between PUFAs, Piezo1 expression, and microglia behavior in the AD brain. These findings could open new possibilities in harnessing microglia in AD and in developing novel therapeutic strategies.

From chronic stress and anxiety to neurodegeneration: Focus on neuromodulation of the axon initial segment.

To adapt to the sustained demands of chronic stress, discrete brain circuits undergo structural and functional changes often resulting in anxiety disorders. In some individuals, anxiety disorders precede the development of motor symptoms of Parkinson's disease (PD) caused by degeneration of neurons in the substantia nigra (SN). Here, we present a circuit framework for probing a causal link between chronic stress, anxiety, and PD, which postulates a central role of abnormal neuromodulation of the SN's axon initial segment by brainstem inputs. It is grounded in findings demonstrating that the earliest PD pathologies occur in the stress-responsive, emotion regulation network of the brainstem, which provides the SN with dense aminergic and cholinergic innervation. SN's axon initial segment (AIS) has unique features that support the sustained and bidirectional propagation of activity in response to synaptic inputs. It is therefore, especially sensitive to circuit-mediated stress-induced imbalance of neuromodulation, and thus a plausible initiating site of neurodegeneration. This could explain why, although secondary to pathophysiologies in other brainstem nuclei, SN degeneration is the most extensive. Consequently, the cardinal symptom of PD, severe motor deficits, arise from degeneration of the nigrostriatal pathway rather than other brainstem nuclei. Understanding when and how circuit dysfunctions underlying anxiety can progress to neurodegeneration, raises the prospect of timed interventions for reversing, or at least impeding, the early pathophysiologies that lead to PD and possibly other neurodegenerative disorders.

Short-term fish oil supplementation applied in presymptomatic stage of Alzheimer's disease enhances microglial/macrophage barrier and prevents neuritic dystrophy in parietal cortex of 5xFAD mouse model.

Dystrophic neurites and activated microglia are one of the main neuropathological characteristics of Alzheimer's disease (AD). Although the use of supplements with omega-3 fatty acids has been associated with reduced risk and lessened AD pathology, it still remains elusive whether such a treatment could affect dystrophic neurites (DNs) formation and microglia/macrophage behavior in the early phase of disease. We analyzed the effects of short-term (3 weeks) fish oil supplementation on DNs formation, tau hyperphosphorylation, Amyloid-beta peptide 1-42 (Aß42) levels and microglial/macrophage response to AD pathology in the parietal cortex of 4-month-old 5xFAD mice, a mouse model of AD. The present study shows for the first time that short-term FO supplementation applied in presymptomatic stage of AD, alters the behaviour of microglia/macrophages prompting them to establish a physical barrier around amyloid plaques. This barrier significantly suppresses DNs formation through the reduction of both Aß content and tau hyperphosphorylation. Moreover, the short-term FO treatment neither suppresses inflammation nor enhances phagocytic properties of microglia/macrophages in the response to Aß pathology, the effects most commonly attributed to the fish oil supplementation. Our findings suggest that fish oil consumption may play an important role in modulating microglial/macrophage response and ameliorating the AD pathology in presymptomatic stage of Alzheimer's disease.

Short-Term Fish Oil Treatment Changes the Composition of Phospholipids While Not Affecting the Expression of Mfsd2a Omega-3 Transporter in the Brain and Liver of the 5xFAD Mouse Model of Alzheimer's Disease.

Long-term fish oil (FO) supplementation is able to improve Alzheimer's disease (AD) pathology. We aimed to determine the impact of short-term fish oil (FO) intake on phospholipids composition and plaque pathology in 5xFAD mice, a widely used animal model of AD. A 3-week-long FO supplementation administered at 3 months of age decreased the number of dense core plaques in the 5xFAD cortex and changed phospholipids in the livers and brains of wild-type (Wt) and 5xFAD mice. Livers of both genotypes responded by increase of n-3 and reciprocal decrease of n-6 fatty acids. In Wt brains, FO supplementation induced elevation of n-3 fatty acids and subsequent enhancement of n-6/n-3 ratio. However, in 5xFAD brains the improved n-6/n-3 ratio was mainly due to FO-induced decrease in arachidonic and adrenic n-6 fatty acids. Also, brain and liver abundance of n-3 fatty acids were strongly correlated in Wts, oppositely to 5xFADs where significant brain-liver correlation exists only for n-6 fatty acids. Expression of omega-3 transporter Mfs2a remained unchanged after FO supplementation. We have demonstrated that even a short-term FO intake improves the phospholipid composition and has a significant effect on plaque burden in 5xFAD brains when applied in early stages of AD pathology.

Limited daily feeding and intermittent feeding have different effects on regional brain energy homeostasis during aging.

Albeit aging is an inevitable process, the rate of aging is susceptible to modifications. Dietary restriction (DR) is a vigorous nongenetic and nonpharmacological intervention that is known to delay aging and increase healthspan in diverse species. This study aimed to compare the impact of different restricting feeding regimes such as limited daily feeding (LDF, 60% AL) and intermittent feeding (IF) on brain energy homeostasis during aging. The analysis was focused on the key molecules in glucose and cholesterol metabolism in the cortex and hippocampus of middle-aged (12-month-old) and aged (24-month-old) male Wistar rats. We measured the impact of different DRs on the expression levels of AMPK, glucose transporters (GLUT1, GLUT3, GLUT4), and the rate-limiting enzyme in the cholesterol synthesis pathway (HMGCR). Additionally, we assessed the changes in the amounts of cholesterol, its metabolite, and precursors following LDF and IF. IF decreased the levels of AMPK and pAMPK in the cortex while the increased levels were detected in the hippocampus. Glucose metabolism was more affected in the cortex, while cholesterol metabolism was more influenced in the hippocampus. Overall, the hippocampus was more resilient to the DRs, with fewer changes compared to the cortex. We showed that LDF and IF differently affected the brain energy homeostasis during aging and that specific brain regions exhibited distinct vulnerabilities towards DRs. Consequently, special attention should be paid to the DR application among elderly as different phases of aging do not respond equally to altered nutritional regimes.

Cholesterol metabolism changes under long-term dietary restrictions while the cholesterol homeostasis remains unaffected in the cortex and hippocampus of aging rats.

Maintaining cholesterol homeostasis in the brain is vital for its proper functioning. While it is well documented that dietary restriction modulates the metabolism of cholesterol peripherally, little is known as to how it can affect cholesterol metabolism in the brain. The present study was designed to elucidate the impact of long-term dietary restriction on brain cholesterol metabolism. Three-month-old male Wistar rats were exposed to long-term dietary restriction until 12 and 24 months of age. The concentrations of cholesterol, its precursors and metabolites, and food-derived phytosterols were measured in the serum, cortex, and hippocampus by gas chromatography/mass spectrometry. Relative changes in the levels of proteins involved in cholesterol synthesis, transport, and degradation were determined by Western blot analysis. Reduced food intake influenced the expression patterns of proteins implicated in cholesterol metabolism in the brain in a region-specific manner. Dietary restriction decreased the concentrations of cholesterol precursors, lanosterol in the cortex, and lanosterol and lathosterol in the hippocampus at 12 months, while the level of desmosterol was elevated in the hippocampus at 24 months. The concentrations of cholesterol and 24(S)-hydroxycholesterol remained unaffected. Food-derived phytosterols were significantly lower after dietary restriction in both the cortex and hippocampus at 12 and 24 months. These findings provide new insight into the effects of dietary restriction on cholesterol metabolism in the brain, lending further support to its neuroprotective effect.

Direct inhibition of myosin II effectively blocks glioma invasion in the presence of multiple motogens.

Anaplastic gliomas, the most common and malignant of primary brain tumors, frequently contain activating mutations and amplifications in promigratory signal transduction pathways. However, targeting these pathways with individual signal transduction inhibitors does not appreciably reduce tumor invasion, because these pathways are redundant; blockade of any one pathway can be overcome by stimulation of another. This implies that a more effective approach would be to target a component at which these pathways converge. In this study, we have investigated whether the molecular motor myosin II represents such a target by examining glioma invasion in a series of increasingly complex models that are sensitive to platelet-derived growth factor, epidermal growth factor, or both. Our results lead to two conclusions. First, malignant glioma cells are stimulated to invade brain through the activation of multiple signaling cascades not accounted for in simple in vitro assays. Second, even though there is a high degree of redundancy in promigratory signaling cascades in gliomas, blocking tumor invasion by directly targeting myosin II remains effective. Our results thus support our hypothesis that myosin II represents a point of convergence for signal transduction pathways that drive glioma invasion and that its inhibition cannot be overcome by other motility mechanisms.

Constitutive EGFR signaling in oligodendrocyte progenitors leads to diffuse hyperplasia in postnatal white matter.

Gliogenesis requires the careful orchestration of migration, differentiation, and proliferation of progenitors. Signaling through the epidermal growth factor receptor (EGFR) has been implicated in regulating these processes in a variety of cell types, including neural progenitors. By retroviral infection, we constitutively expressed an EGFR-GFP fusion protein in white matter glial progenitors at postnatal day 3 of the rat forebrain in vivo and analyzed the development of these cells over the subsequent 15 weeks. EGFR-GFP+ cells remained proliferative and migratory, gradually populating the brains ipsilateral and contralateral to the side of viral infection, but never differentiated into mature glia. The accumulation of these cells doubled the total cell density in white matter and led to a 10-fold increase in the abundance of glial progenitors, giving rise to a progenitor "hyperplasia." The marker profile of infected cells, NG2+, olig2+, PDGFR-alpha+, nestin+, GFAP-, and CC1-, indicated a close resemblance to oligodendrocyte progenitors. Positive immunostaining for phosphorylated EGFR colocalized with punctate accumulation of EGFR-GFP, indicating that a subset of receptors was engaged in active signaling. Furthermore, EGF was required to observe phospho-tyrosine EGFR immunostaining of glial progenitors in culture. These observations suggest that constitutive EGFR expression can inhibit glial differentiation, but requires ligand as well.

DARPP-32 genomic fragments drive Cre expression in postnatal striatum.

To direct Cre-mediated recombination to differentiated medium-size spiny neurons (MSNs) of the striatum, we generated transgenic mice that express Cre recombinase under the regulation of DARPP-32 genomic fragments. In this reported line, recombination of an R26R reporter allele occurred postnatally in the majority of medium-size spiny neurons of the dorsal and ventral striatum (caudate nucleus and nucleus accumbens/olfactory tubercle), as well as in the piriform cortex and choroid plexus. Although regulatory fragments were selected to target MSNs, low levels of Cre-recombinase expression, as detected by beta-galactosidase activity from the R26R reporter gene, were also apparent in widely dispersed areas or cells of the forebrain and hindbrain. These included the primary and secondary motor cortex, and association cortex, as well as in the olfactory bulb and cerebellar Purkinje cells. Notably, expression in these regions was well below that of endogenous DARPP-32. Analysis of colocalization of beta-galactosidase, as detected either by histochemistry or immunocytochemistry, and DARPP-32 revealed double-labeling in almost all DARPP-32-expressing MSNs in the postnatal striatum, but not in extrastriatal regions. The DARPP-32Cre transgenic mouse line thus provides a useful tool to specifically express and/or inactivate genes in mature MSNs of the striatum.