Cystatin C prevents neuronal loss and behavioral deficits via the endosomal pathway in a mouse model of down syndrome.

Cystatin C (CysC) plays diverse protective roles under conditions of neuronal challenge. We investigated whether CysC protects from trisomy-induced pathologies in a mouse model of Down syndrome (DS), the most common cause of developmental cognitive and behavioral impairments in humans. We have previously shown that the segmental trisomy mouse model, Ts[Rb(12.1716)]2Cje (Ts2) has DS-like neuronal and behavioral deficiencies. The current study reveals that transgene-mediated low levels of human CysC overexpression has a preventive effect on numerous neuropathologies in the brains of Ts2 mice, including reducing early and late endosome enlargement in cortical neurons and decreasing loss of basal forebrain cholinergic neurons (BFCNs). Consistent with these cellular benefits, behavioral dysfunctions were also prevented, including deficits in nesting behavior and spatial memory. We determined that the CysC-induced neuroprotective mechanism involves activation of the phosphotidylinositol kinase (PI3K)/AKT pathway. Activating this pathway leads to enhanced clearance of accumulated endosomal substrates, protecting cells from DS-mediated dysfunctions in the endosomal system and, for BFCNs, from neurodegeneration. Our findings suggest that modulation of the PI3/AKT pathway offers novel therapeutic interventions for patients with DS.

Glutamatergic transmission aberration: a major cause of behavioral deficits in a murine model of Down's syndrome.

Trisomy 21, or Down's syndrome (DS), is the most common genetic cause of intellectual disability. Altered neurotransmission in the brains of DS patients leads to hippocampus-dependent learning and memory deficiency. Although genetic mouse models have provided important insights into the genes and mechanisms responsible for DS-specific changes, the molecular mechanisms leading to memory deficits are not clear. We investigated whether the segmental trisomy model of DS, Ts[Rb(12.1716)]2Cje (Ts2), exhibits hippocampal glutamatergic transmission abnormalities and whether these alterations cause behavioral deficits. Behavioral assays demonstrated that Ts2 mice display a deficit in nest building behavior, a measure of hippocampus-dependent nonlearned behavior, as well as dysfunctional hippocampus-dependent spatial memory tested in the object-placement and the Y-maze spontaneous alternation tasks. Magnetic resonance spectra measured in the hippocampi revealed a significantly lower glutamate concentration in Ts2 as compared with normal disomic (2N) littermates. The glutamate deficit accompanied hippocampal NMDA receptor1 (NMDA-R1) mRNA and protein expression level downregulation in Ts2 compared with 2N mice. In concert with these alterations, paired-pulse analyses suggested enhanced synaptic inhibition and/or lack of facilitation in the dentate gyrus of Ts2 compared with 2N mice. Ts2 mice also exhibited disrupted synaptic plasticity in slice recordings of the hippocampal CA1 region. Collectively, these findings imply that deficits in glutamate and NMDA-R1 may be responsible for impairments in synaptic plasticity in the hippocampus associated with behavioral dysfunctions in Ts2 mice. Thus, these findings suggest that glutamatergic deficits have a significant role in causing intellectual disabilities in DS.

Sphenoid Sinus Aspergilloma in an Immunocompetent and an Immunocompromised Patient: A Case Report.

Sellar, supra-sellar aspergilloma are rare differentials for a sellar mass. CNS aspergilloma occurs due to intracranial extension of invasive fungal sinusitis, and often first manifests with symptoms of headache and visual disturbance. This complication is much more common in immunocompromised patients, but proliferation of fungal pathogens and low index for suspicion has led to more severe breakthrough cases in the immunocompetent. If treated timely, these CNS lesions can have a relatively good prognosis. Conversely, delays in diagnosis can confer very high rates of mortality among patients with invasive fungal disease. Originally from India, in this case report, we describe two patients presenting with sellar, supra-sellar tumors, who eventually were diagnosed with confirmed cases of invasive intracranial aspergilloma. We describe the clinical presentation, imaging techniques, and treatment modalities for this relatively rare disease in both the immunocompromised and the immunocompetent.

Perioperative Management of Spontaneous Intracranial Hemorrhage in a Patient With Hemophilia A in a Resource Limited Country.

Intracranial hemorrhage (ICH) is a serious complication of hemophilia A with high morbidity and mortality. The management of such cases is complicated by nonspecific and often delayed presentation, increased frequency of rebleeding, low awareness regarding clotting factor replacement, and debate regarding the efficacy of surgical interventions. We report a case of an 18-year-old male patient with hemophilia A, who first presented to the emergency department in India in a comatose state. Neuroimaging revealed subdural hematoma with midline shift and uncal herniation. The patient was successfully managed with perioperative cryoprecipitate and factor VIII replacement, tiered intracranial pressure lowering strategies, and early decompressive craniectomy with clot evacuation. In India, there are no standardized guidelines for screening and routine care for hereditary diseases like hemophilia. In a resource-deficient country, management was complicated by the limited availability of factor VIII in the emergent setting, as well as the inability to obtain serial factor levels in the postoperative period. We hope that this article helps to guide the management of ICH and hemophilia in resource-limited countries.

Automatic Identification of Glomerular in Whole-Slide Images Using a Modified UNet Model.

Glomeruli are interconnected capillaries in the renal cortex that are responsible for blood filtration. Damage to these glomeruli often signifies the presence of kidney disorders like glomerulonephritis and glomerulosclerosis, which can ultimately lead to chronic kidney disease and kidney failure. The timely detection of such conditions is essential for effective treatment. This paper proposes a modified UNet model to accurately detect glomeruli in whole-slide images of kidney tissue. The UNet model was modified by changing the number of filters and feature map dimensions from the first to the last layer to enhance the model's capacity for feature extraction. Moreover, the depth of the UNet model was also improved by adding one more convolution block to both the encoder and decoder sections. The dataset used in the study comprised 20 large whole-side images. Due to their large size, the images were cropped into 512 × 512-pixel patches, resulting in a dataset comprising 50,486 images. The proposed model performed well, with 95.7% accuracy, 97.2% precision, 96.4% recall, and 96.7% F1-score. These results demonstrate the proposed model's superior performance compared to the original UNet model, the UNet model with EfficientNetb3, and the current state-of-the-art. Based on these experimental findings, it has been determined that the proposed model accurately identifies glomeruli in extracted kidney patches.

Documenting Disease in the Undocumented Migrants: A Case Report of Chronic Kidney Disease of Unknown Origin in a Central American Migrant.

Mesoamerican nephropathy (MeN) or chronic kidney disease of unknown origin (CKDu) is a rising epidemic in hotspot regions of El Salvador and Nicaragua. MeN is often defined in patients who exhibit a clinically reduced estimated glomerular filtration rate (eGFR) but lack a defining etiology such as diabetes or hypertension. A multitude of risk factors for MeN have been identified, including physical labor demands in a hot climate, exposure to pesticides, and poverty. Additionally, social determinants such as limited access to health care and the cost of disease burden often contribute to overall poor prognosis and progression of the disease. We present a case of a 39-year-old male with a past medical history of gout who presented to the emergency room with abdominal pain radiating to the flanks and bilateral great toe pain. Social history revealed the patient recently moved to the United States from Central America (Nicaragua), was unemployed, and did not have health insurance. Prior to the presentation, the patient admitted he was not compliant with his gout medications for about one month. The symptoms first began two to three weeks prior to his evaluation in the emergency department; the patient also endorsed decreased oral intake during this time period. He was noted to have abnormally elevated creatinine along with elevated uric acid levels, low potassium and magnesium levels. Abdominal imaging revealed nephrolithiasis without hydronephrosis. Initial differentials included acute kidney injury (AKI) from dehydration, non-steroidal anti-inflammatory drug (NSAID) induced nephropathy, and uric acid nephropathy. This patient was eventually found to have a biopsy-proven findings of CKDu. We want to highlight the need to keep MeN high in the differential with a low threshold to perform a renal biopsy for accurate diagnosis and management of the disease, especially in the rising immigrant population in the United States.

The Linkage Between Autism Spectrum Disorder and Dup15q Syndrome: A Case Report.

Patients diagnosed with autism spectrum disorder (ASD) frequently have a variable presentation and can suffer from underlying conditions, such as Chromosome 15 abnormalities] The broad diagnosis of ASD and its debilitating symptoms can overshadow underlying conditions and delay crucial interventions. This report describes a male child who was diagnosed with ASD at the early age of 19 months. Hallmark symptoms seen in this case included lack of social eye contact, lack of joint attention, hand-flapping, and missed motor milestones. Genetic methylation assay revealed a duplication on maternally derived chromosome 15, indicating concurrent 15q11-q13 duplication syndrome (Dup15q). Screening assessments for ASD are an important step in the initial management of developmental abnormalities. However, early genetic screening can lead to a more accurate diagnosis, personalized treatment, and better quality of life in patients with atypical symptoms caused by undiagnosed comorbid conditions.

Advanced non-small cell lung cancer treated with palliative systemic therapy complicated by calvarial metastasis: a case report.

Background: Bony metastases are often seen in advanced cancers and lead to deterioration in patient quality of life with common complications of pain, bone fractures, and hypercalcemia. While most sites of metastasis to bone are observed in the axial skeleton from patients with a primary lung, breast or prostate cancer, metastases to the calvarium from lung cancer are less common, and thus less likely to be identified and managed. Case Description: A 69-year-old Caucasian female with advanced non-small cell lung cancer (NSCLC) presented with worsening symptoms of widespread body pain, fatigue, and weight loss. Physical examination was remarkable for a palpable protrusion on the patient's head. Imaging revealed a parieto-occipital calvarial lesion, a likely metastasis from her lung cancer. A previously performed CT-guided lung biopsy was evaluated for actionable tumor markers to allow for more specific and efficacious line of treatments; the patient's tumor had lacked any notable gene mutations. The treatment plan included radiotherapy, combined immunotherapy and chemotherapy consisting of pembrolizumab, pemetrexed, and carboplatin. Despite the treatment, the patient's skull lesion had continued to grow, and her overall condition deteriorated to the point where she required hospice. Conclusions: Given the unique location of calvarial metastases, early detection appears to correlate with improving patient outcomes and quality of life. A multimodal approach with a high index of suspicion is essential for diagnosing and managing rare presentations of metastatic disease.

Unfolding of Novel Independent Missense Mutations in VAMP2 and AGRN and Their Collective Role in Global Developmental Delay: A Case Report.

Vesicle-associated membrane protein 2 (VAMP2) and Agrin (AGRN) are crucial proteins in neurotransmission. VAMP2 is a vesicular protein that facilitates the exocytosis of neurotransmitters. At the same time, AGRN plays a critical role in the maintenance and function of neuromuscular junctions. Mutations in the signaling pathway of VAMP2 and AGRN impair proper signaling between the presynaptic and postsynaptic neurons, and can result in neurodevelopmental conditions known as global developmental delay (GDD). This study highlights a presentation of GDD in a patient with concurrent mutations in VAMP2 and AGRN. A three-year-old female child presented with GDD characterized by hypotonia, intellectual disability, and dysphagia. Physical exam exhibited signs of developmental delay and severe muscle weakness. EEG findings were suggestive of a hypsarrhythmia pattern. The ophthalmological evaluation showed partial optic atrophy bilaterally. Therapeutic interventions included Keppra and Topamax, which proved ineffective. The patient's outcome was inconclusive as care was transferred to another facility. This case study reports the novel appearance of two concurrent mutations: p.Gln76Pro associated with VAMP2 and p.Gln970Glu associated with AGRN. Mutations in VAMP2 lead to a dysfunctional SNARE complex and inhibit exocytosis of neurotransmitters into the synaptic cleft. Mutations in AGRN impair the ability to form and activate postsynaptic nicotinic acetylcholine receptors. Improper signaling between presynaptic and postsynaptic neurons is an important determinant of GDD. We hope that accounting for this mutational pattern will contribute to understanding synapse assembly and help unravel the complex interplay of factors involved in the pathology of neuromuscular disorders and GDD.

Cystatin C rescues degenerating neurons in a cystatin B-knockout mouse model of progressive myoclonus epilepsy.

In vitro studies have shown that cystatin C (CysC) is neuroprotective. Here we demonstrate that CysC is neuroprotective in vivo, in a mouse model of the inherited neurodegenerative disorder, progressive myoclonic epilepsy type 1 (EPM1). Loss-of-function mutations in the cystatin B (CysB) gene, an intracellular cysteine protease inhibitor, lead to this human disease. A CysB-knockout (CysBKO) mouse model develops symptoms that mimic EPM1. CysB deficiency in these mice results in enhanced cathepsin B and D activities, indicating lysosomal dysfunction. We show that expression of CysC is enhanced in the brains of CysBKO mice. Crossbreeding of CysBKO mice with either CysC-overexpressing transgenic mice or CysC-knockout mice demonstrates that clinical symptoms and neuropathologies, including motor coordination disorder, cerebellar atrophy, neuronal loss in the cerebellum and cerebral cortex, and gliosis caused by CysB deficiency, are rescued by CysC overexpression and exacerbated by CysC deficiency. Thus, CysC effectively rescues the CysB loss-of-function mutations, facilitating the reversal of pathophysiological changes and suggesting a novel therapeutic intervention for patients with EPM1 and other neurodegenerative disorders.

Combatting the imbalance of sex ratio at birth: medium-term impact of India's National Programme of Beti Bachao Beti Padhao in the Haryana State of India.

The Government of India initiated the Beti Bachao Beti Padhao (B3P) programme in 2015 as a flagship initiative to reduce gender imbalance in sex ratio at birth (SRB) and to ensure social protection of girls. The present study was conducted to evaluate the medium-term impact of B3P implementation in Haryana state, from 2015 to 2019, on SRB. Monthly data on SRB were collected for the entire state of Haryana through a civil registration system. Segmented time series regression analysis was used to estimate the variations in SRB after the B3P programme with the help of Winter's additive interrupted time series model. The SRB in Haryana increased from 876 girls per 1000 boys in 2015 to 923 in 2019. The results of the model demonstrated that before the inception of intervention (pre-slope), there was a significant monthly change in SRB of 0.217 (95% confidence interval: 0.144-0.290). Following the B3P programme, SRB was found to increase by 0.835 per month, which implied that an increase of 0.618 (confidence interval: 0.338, 0.898) every month in SRB can be attributed to the B3P programme. This indicated that SRB for the state of Haryana increased at the rate of 7.42 units per year as a result of the B3P programme. B3P has led to a significant improvement in SRB in Haryana state. The continuity of efforts in the same direction with a sustained focus on behaviour change will further help achieve the goal of gender parity in births and child survival.

Low-dose ketamine as an adjuvant for pain control in a cancer patient: a case report.

Cancer patients often suffer from pain related problems such as under-treatment of pain, ineffective and persistent opioid administration as well as adverse opioid use outcomes. There is a growing need for non-opioid analgesic alternatives for patients undergoing treatment for obstinate pain. Ketamine is a fast-acting N-methyl-D-aspartate (NMDA) receptor antagonist that has been emerging as an effective medication for pain alleviation. While protocols have been established for the use of Low-Dose Ketamine (LDK) for post-operative pain, there is growing evidence for using LDK as a clinical alternative to opioids in a palliative care setting. This case study involves monitoring the efficacy of LDK treatment in combination with opioid analgesics in a cancer patient in a hospital setting. This is a very selected case of a patient with Metastatic Prostate Cancer (Gleason 9 Adenocarcinoma) where LDK was shown to be efficacious at reducing pain when opioids and standard pain medications were not satisfactory. While the study involved using a relatively novel pharmacological protocol and close patient monitoring, the patient reported a sustained reduction in pain level based on the Numerical Rating Scale for months after the termination of LDK infusions. Moreover, the treatment also resulted in a reduction of total opioid usage after the addition of LDK. Although additional research is needed to ascertain optimal dosing schedules and route of Ketamine, given these promising findings, Ketamine may be a useful option for improving the treatment of refractory pain in patients with cancer and a good tool in palliative medicine for treating neoplastic pain.

Low-Temperature Chemical Vapor Deposition Growth of Graphene Layers on Copper Substrate Using Camphor Precursor.

A two-step, low-temperature thermal chemical vapor deposition (CVD) process, which uses camphor for synthesizing continuous graphene layer on Cu substrate is reported. The growth process was performed at lower temperature (800 °C) using camphor as the source of carbon. A threezone CVD system was used for controlled heating of precursor, in order to obtain uniform graphene layer. As-grown samples were characterized by X-ray diffraction (XRD), Raman spectroscopy and transmission electron microscopy (TEM). The results show the presence of 4-5 layers of graphene. As-grown graphene transferred onto a glass substrate through a polymer-free wet-etching process, demonstrated transmittance ~91% in visible spectra. This process of synthesizing large area, 4-5 layer graphene at reduced temperature represents an energy-efficient method of producing graphene for possible applications in opto-electronic industry.

Distinct Levels of Adhesion Energy of In-Situ Grown CuO Nanostructures.

CuO nanostructures were reported for a myriad of applications in diverse areas such as high Tc superconductors, field emitters, catalysts, gas sensors, magnetic storage, biosensors, superhydrophobic surfaces, energy materials etc. In all these applications, structural stability of the nanostructures is very important for efficient functioning of devices with a longer lifetime. Hence, it is necessary to understand the adhesion energy of these nanostructures with their substrates. In this research work, a variety of CuO nanostructures were synthesized directly on Cu foil substrate by varying only the concentration of the reagents. CuO nanostructures, thus grown, were subjected to a nano-scratch test to quantify their adhesion strength with Cu substrate. The adhesion energy was observed to be highest for nanorods and lowest for nanoribbons among all the CuO nanostructures synthesized in this work. Results of this research will be useful in predicting the service life and in improving the efficiency of CuO nanostructure-based devices.

Association of vitamin B12 mediated hyperhomocysteinemia with depression and anxiety disorder: A cross-sectional study among Bhil indigenous population of India.

BACKGROUND: Indigenous populations in India are amongst the poorest and most marginalized population groups experiencing severe health deprivation. AIM: The present study is the first study that aims to understand the association of micronutrient deficiencies (vitamin B12 and folate) and MTHFR C677T gene polymorphism with depression and generalized anxiety disorder (GAD) among the Bhil indigenous population of India. METHODS: A total of 303 participants aged 25-65 years of both sexes and unrelated up to first cousins belonging to Bhil indigenous population were recruited for the present study. Depression and generalized anxiety disorder were assessed using Patient Health Questionnaire and Generalized Anxiety Disorder scale, respectively. Biochemical analysis, DNA extraction and MTHFR C677T gene polymorphism analysis were done using standard protocols. RESULTS: Although, vitamin B12 and folate status was not found to be directly associated with depression and GAD, but hyperhomocysteinemia was posing more than three folds and six folds significant increased risk for depression and GAD, respectively. Further, it seems hyperhomocysteinemia was mediated by vitamin B12 deficiency among depressed and anxious individuals. T allele of MTHFR C677T gene polymorphism was posing increased risk for depression and anxiety disorder though not significant. CONCLUSION: The present study highlights the significance of micronutrient deficiencies in the causation of depression and anxiety disorder.

A pleiotropic role for exosomes loaded with the amyloid ß precursor protein carboxyl-terminal fragments in the brain of Down syndrome patients.

Down syndrome (DS) is characterized by cognitive deficits throughout the life span and with the development of aging-dependent Alzheimer's type neuropathology, which is related to the triplication of the amyloid ß precursor protein (APP) gene. A dysfunctional endosomal system in neurons is an early characteristic of DS and APP metabolites accumulate in endosomes in DS neurons. We have previously shown enhanced release of exosomes in the brain of DS patients and the mouse model of DS Ts[Rb(12.1716)]2Cje (Ts2), and by DS fibroblasts, as compared with diploid controls. Here, we demonstrate that exosome-enriched extracellular vesicles (hereafter called EVs) isolated from DS and Ts2 brains, and from the culture media of human DS fibroblasts are enriched in APP carboxyl-terminal fragments (APP-CTFs) as compared with diploid controls. Moreover, APP-CTFs levels increase in an age-dependent manner in EVs isolated from the brain of Ts2 mice. The release of APP-CTFs-enriched exosomes may have a pathogenic role by transporting APP-CTFs into naïve neurons and propagating these neurotoxic metabolites, which are also a source of amyloid ß, throughout the brain, but also provides a benefit to DS neurons by shedding APP-CTFs accumulated intracellularly.

Characterizing molecular probes for diffusion measurements in the brain.

Brain diffusion properties are at present most commonly evaluated by magnetic resonance (MR) diffusion imaging. MR cannot easily distinguish between the extracellular and intracellular signal components, but the older technique of real-time iontophoresis (RTI) detects exclusively extracellular diffusion. Interpretation of the MR results would therefore benefit from auxiliary RTI measurements. This requires a molecular probe detectable by both techniques. Our aim was to specify a minimum set of requirements that such a diffusion probe should fulfill and apply it to two candidate probes: the cation tetramethylammonium (TMA(+)), used routinely in the RTI experiments, and the anion hexafluoroantimonate (SbF(6)(-)). Desirable characteristics of a molecular diffusion probe include predictable diffusion properties, stability, minimum interaction with cellular physiology, very slow penetration into the cells, and sufficiently strong and selective MR and RTI signals. These properties were evaluated using preparations of rat neocortical slices under normal and ischemic conditions, as well as solutions and agarose gel. While both molecules can be detected by MR and RTI, neither proved an ideal candidate. TMA(+) was very stable but it penetrated into the cells and accumulated there within tens of minutes. SbF(6)(-) did not enter the cells as readily but it was not stable, particularly in ischemic tissue and at higher temperatures. Its presence also resulted in a decreased extracellular volume. These probe properties help to interpret previously published MR data on TMA(+) diffusion and might play a role in other diffusion experiments obtained with them.

Blunted amygdalar anti-inflammatory cytokine effector response to postnatal stress in prenatally stressed rats.

Smad1 is the downstream effector for bone morphogenetic protein, part of the anti-inflammatory cytokine family. Glucocorticoids (GCs) increase the production of anti-inflammatory cytokines to oppose the actions of pro-inflammatory cytokines. Here we used the prenatally stressed (PS) rat to see if chronic GC activation affects this protective mechanism in the amygdala. Male PS and control offspring were either left undisturbed or exposed to a 2-week regimen of intruder stress. One week later, half of these animals were further subjected to restraint stress for 3 days. Nuclear and cytoplasmic phosphorylated (p)-Smad1 were visualized by immunocytochemistry and quantified in the lateral and basolateral amygdala and in the hind limb primary somatosensory (S1HL) cortex. PS rats showed significantly greater baseline p-Smad1 per cell than controls. However, intruder stress increased p-Smad1 nuclear staining in the control rats only: no further increases in either compartment were observed in the PS group. With repeated restraint stress, attenuation of both cytoplasmic and nuclear p-Smad1 responses was significantly greater in controls. Thus, the overall p-Smad1 responsiveness of amygdala neurons of PS rats to life stressors is blunted. We hypothesize that the amygdala may play an essential role in initiating the cytokine response to stress in the adult rat brain. Basal p-Smad1 staining was unaffected by prenatal stress in the S1HL cortex but became elevated in the cytoplasm following intruder stress. The significance of this is unknown, but may point to a means by which stress can generally affect cells whose functions are unrelated to driving the sympathoadrenal system.

Association of vitamin B12 mediated hyperhomocysteinemia and methylenetetrafolate reductase (C677T) gene polymorphism with cognitive impairment: A population based study from North India.

The present study attempts to understand the association of homocysteine, vitamin B12, folate, and MTHFR C677T gene polymorphism with cognitive impairment (CI) among 808 individuals of either sex (aged 30-70 years) from a largely vegetarian, mendelian population of North India. Biochemical and genetic analyses were done using standard protocols. Results indicate that 34.3% of the subjects had mild CI, 28.7% moderate CI and 0.2% were having severe CI. Hyperhomocysteinemia was found to be a significant risk factor for moderate/severe CI. Both CT genotype and T allele of MTHFR C677T gene polymorphism were found to pose significant decreased risk for CI.

Estrogen regulation of neurotrophin expression in sympathetic neurons and vascular targets.

We hypothesize that estrogen exerts a modulatory effect on sympathetic neurons to reduce neural cardiovascular tone and that these effects are modulated by nerve growth factor (NGF), a neurotrophin that regulates sympathetic neuron survival and maintenance. We examined the effects of estrogen on NGF and tyrosine hydroxylase (TH) protein content in specific vascular targets. Ovariectomized, adult Sprague-Dawley rats were implanted with placebo or 17beta-estradiol (release rate, 0.05 mg/day). Fourteen days later, NGF levels in the superior cervical ganglia (SCG) and its targets, the heart, external carotid artery, and the extracerebral blood vessels, as well as estrogen receptor alpha (ERalpha) content levels in the heart, were determined using semi-quantitative Western blot analysis. TH levels in the SCG and extracerebral blood vessels were determined by Western blotting and immunocytochemistry, respectively. Circulating levels of 17beta-estradiol and prolactin (PRL) were quantified by RIA. Estrogen replacement significantly decreased NGF protein in the SCG and its targets, the external carotid artery, heart and extracerebral blood vessels. TH protein associated with the extracerebral blood vessels was also significantly decreased, but ERalpha levels were significantly increased in the heart following estrogen replacement. These results indicate that estrogen reduces NGF protein content in sympathetic vascular targets, which may lead to decreased sympathetic innervations to these targets, and therefore reduced sympathetic regulation. In addition, the estrogen-induced increase in ERalpha levels in the heart, a target tissue of the SCG, suggests that estrogen may sensitize the heart to further estrogen modulation, and possibly increase vasodilation of the coronary vasculature.