Puberty is the attainment of fertility -- the ability to reproduce. It occurs because our brains ... more Puberty is the attainment of fertility -- the ability to reproduce. It occurs because our brains begin to secrete one key hormone, gonadotropin-releasing hormone. This causes the pituitary gland to induce maturation of the testis and the ovary, which then produce sex steroid hormones and mature sperm and eggs. We should be concerned that the age at which puberty begins is decreasing in many countries. Puberty starts when adequate growth and energy storage has occurred, so the earlier age of puberty may reflect recent increases in adolescent obesity resulting from more sedentary lifestyles.
bioRxiv (Cold Spring Harbor Laboratory), Dec 16, 2021
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycytespecific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylasepositive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Singlecell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced expression in the mutant mouse. However, the mild tanycyte depletion and loss of markers observed in NrCAM-deficient mice were associated with only a subtle metabolic phenotype.
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Single-cell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced express...
Twelve lactating sows were used at 22.4±0.8 days postpartum to determine whether endogenous opioi... more Twelve lactating sows were used at 22.4±0.8 days postpartum to determine whether endogenous opioid peptides (FOP) are involved in the suckling-induced inhibition of luteinizing hormone (LH) secretion. Four sows each received either 1, 2, or 4 mg/kg body weight of naloxone (NA L), an opiate antagonist, in saline i. v. Blood was collected at 15-mm intervals for 2 h before and 4 b after NAL treatment. All sows were then given 100 pg gonadotropin-releasing hormone (GnRH) in saline iv., and blood samples were collected for an additional h. Pigs were weaned after blood sampling. At 40 b after weaning, sows were treated and blood samples collected as during suckling. Serum concentrations of LH after treatment with NAL were similar for all doses; therefore, the data were pooled across doses. During suckling, serum concentrations of LH were 0.41±0.04 ng/ml before NAL treatment, increased to 0.65±0.08 ng/ml at 30 mm after NAL treatment, and remained elevated above pretreatmerit concentrations for 120 mm (p<0.05). Naloxone failed to alter serum concentrations of LH after weaning. Tbese data indicate that EOP may be involved in the suckling-induced suppression of LH secretion and that weaning may either decrease opioid inhibition of LH secretion or decrease pituitary LH responsiveness to endogenous GnRH released by NAL.
4-£ F x X. rt E. S' 3 I? 5' ... Melatonin and the Mammalian Pineal Gland This One 66C3... more 4-£ F x X. rt E. S' 3 I? 5' ... Melatonin and the Mammalian Pineal Gland This One 66C3-RWT-826F ... Melatonin and the Mammalian Pineal Gland Josephine Arendt Professor of Endocrinology School of Biological Sciences University of Surrey Guildford, UK ...
Background and aimTherapeutic activation of thermogenic brown adipose tissue (BAT) is a potential... more Background and aimTherapeutic activation of thermogenic brown adipose tissue (BAT) is a potential strategy to prevent obesity and metabolic disease in humans. However, it is now recognised that rodent studies examining BAT physiology are carried out at sub‐thermoneutral temperatures (e.g. ~20°C), and are not translationally relevant to humans as BAT is ‘hyperactive’. Therefore, the aim of this study was to determine the effect of common regulators of BAT metabolism when animals were raised at thermoneutrality (28°C).Material and methodsThirty weanling Sprague‐Dawley rats were housed at thermoneutrality (28°C) and randomised to either chow (C, n=6) or a high‐fat diet (HFD, n=24) from 3‐weeks of age. At 12 weeks, subgroups (n=6) of HFD were randomised to either mild‐cold exposure (20°C), Mirabegron, a selective β3‐agonist (0.75 mk/kg/d) or exercise training (1h/d, 5 d/week). Metabolic assessment was undertaken in CLAMS during the last 48h to assess energy intake (EI), expenditure (EE) and physical activity (PA) in addition to the acute response to administration of Mirabegron. Key thermogenic and metabolic genes were analysed in interscapular BAT by qPCR in addition to targeted insulin resistance PCR Arrays (86 key genes, n=3).ResultsNo interventions reduced body weight or fat mass. Intriguingly however, mild‐cold exposure significantly increased weight‐gain during the 4‐week period (78.6 vs. 119.8g). This was accompanied by a significant increase in inguinal AT (7.14 vs 16.14g) in cold‐exposed animals whilst BAT mass was significantly increased with exercise‐training (0.74 vs. 1.2g). There was no difference in 24h EE, EI or PA between groups. Key thermogenic genes in BAT were unchanged by the interventions. CITED1 expression was upregulated by HFD and reduced by all interventions whilst PRDM16 expression was reduced by HFD and increased by exercise. Similarly, expression of PPARA, mTOR and the ‘beige’ marker TBX1 were upregulated by exercise only. Targeted PCR arrays demonstrated an upregulation of inflammatory markers e.g. TLR4, EMR1, CASP1 and IL18R1 and a downregulation of metabolic genes e.g. SCD1, FASN, ACACB, HK2 with HFD. β3 increased FASN, whilst IL18R1, IL6 and STAT3 were downregulated along with IRS2, LEP, ADIPOR1, INSR and PIK3R1. Exercise upregulated FASN, SCD1, HK, ACACA, ACACB and PDK2 but NLRP3, IL1β, PYCARD, LPL, IRS2, INSR, FABP4 and CD36 were all downregulated.ConclusionWe show there is no consistent upregulation of BAT as determined by key thermogenic genes in response to common stimuli when examined at thermoneutrality. Effects of interventions on BAT carried out at sub‐thermoneutrality are most likely to be a consequence of chronic mild‐cold stress and are unlikely to be translated to humans.Support or Funding InformationBritish Heart FoundationThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Background and aim: Rodents are commonly housed below thermoneutrality and this exposure to 'cold... more Background and aim: Rodents are commonly housed below thermoneutrality and this exposure to 'cold' (i.e. 20°C) activates thermogenic brown (BAT) and beiging of white adipose tissue. Here, we examined whether a standard housing temperature (i.e. 20°C, a reduction in temperature of ~8°C) or YM-178, a highly-selective β 3-adrenoreceptor agonist, in obese animals raised at thermoneutrality, would impact differently on classical BAT or subcutaneous inguinal (IWAT) beige depots. Methods: Eighteen weanling Sprague-Dawley rats were housed at thermoneutrality (28°C) and fed a high-fat diet. At 12 weeks, 6 animals were randomised to either standard housing temperature (20°C, n=6) or to β 3-AR agonist administration (28°C+β3, 0.75mg/kg/d, n=6) for 4 weeks. Metabolic assessment was undertaken during the final 48h, followed by interscapular, perivascular BAT and IWAT sampling for the analysis of thermogenic genes and the proteome. Results: Exposure to 20°C increased weight gain, BAT and IWAT mass. Proteomic analysis of BAT revealed novel pathways associated with cold-induced weight gain (i.e. histone deacetylation, glycosaminoglycan degradation and glycosphingolipid biosynthesis) whilst β 3adrenoreceptor agonism impacted on proteins involved in skeletal muscle contraction and cell differentiation. IWAT of cold-exposed animals exhibited an enrichment of proteins involved NAD+ binding, plus retinol and tyrosine metabolic pathways whilst β 3-AR agonism downregulated ribosomal and upregulated acute phase response proteins. Conclusion: Following diet-induced obesity at thermoneutrality, exposure to 20°C promotes subcutaneous fat deposition in order to reduce heat loss and defend body temperature. In contrast, chronic administration of β 3-AR agonist has minimal metabolic-related effects on adipose tissue. .
The Siberian hamster (Phodopus sungorus) survives winter by decreasing food intake and catabolizi... more The Siberian hamster (Phodopus sungorus) survives winter by decreasing food intake and catabolizing abdominal fat reserves, resulting in a sustained, profound loss of body weight. Hypothalamic tanycytes are pivotal for this process. In these cells, short-winter photoperiods upregulate deiodinase 3, an enzyme that regulates thyroid hormone availability, and downregulate genes encoding components of retinoic acid (RA) uptake and signaling. The aim of the current studies was to identify mechanisms by which seasonal changes in thyroid hormone and RA signaling from tanycytes might ultimately regulate appetite and energy expenditure. proVGF is one of the most abundant peptides in the mammalian brain, and studies have suggested a role for VGF-derived peptides in the photoperiodic regulation of body weight in the Siberian hamster. In silico studies identified possible thyroid and vitamin D response elements in the VGF promoter. Using the human neuroblastoma SH-SY5Y cell line, we demonstrate...
The viral 2A sequence has become an attractive alternative to the traditional internal ribosomal ... more The viral 2A sequence has become an attractive alternative to the traditional internal ribosomal entry site (IRES) for simultaneous over-expression of two genes and in combination with recombinant adeno-associated viruses (rAAV) has been used to manipulate gene expression in vitro. To develop a rAAV construct in combination with the viral 2A sequence to allow long-term over-expression of the vgf gene and fluorescent marker gene for tracking of the transfected neurones in vivo. Transient transfection of the AAV plasmid containing the vgf gene, viral 2A sequence and eGFP into SH-SY5Y cells resulted in eGFP fluorescence comparable to a commercially available reporter construct. This increase in fluorescent cells was accompanied by an increase in VGF mRNA expression. Infusion of the rAAV vector containing the vgf gene, viral 2A sequence and eGFP resulted in eGFP fluorescence in the hypothalamus of both mice and Siberian hamsters, 32 weeks post infusion. In situ hybridisation confirmed t...
Accurate estimation of the number of ovarian follicles at various stages of development is an imp... more Accurate estimation of the number of ovarian follicles at various stages of development is an important indicator of the process of folliculogenesis in relation to the endocrine signals and paracrine/autocrine mechanisms that control the growth and maturation of the oocytes and their supporting follicular cells. There are 10-fold or greater differences in follicular numbers per ovary at similar ages and/or strains reported in earlier studies using various methods, leading to difficulties with interpretation of ovarian function in control vs experimental conditions. This study describes unbiased, assumption-free stereological methods for quantification of early and growing follicular numbers in the mouse ovary. A fractionator approach was used to sample a defined fraction of histological sections of adult wild-type ovaries. Primordial and primary follicles were counted independently with the optical and physical disector methods. The fractionator/disector methods, which are independe...
We investigated whether histaminergic tone contributes to the seasonal catabolic state in Siberia... more We investigated whether histaminergic tone contributes to the seasonal catabolic state in Siberian hamsters by determining the effect of ablation of histaminergic neurons on food intake, metabolic rate and body weight. A ribosomal toxin (saporin) conjugated to orexin-B was infused into the ventral tuberomammillary region of the hypothalamus, since most histaminergic neurons express orexin receptors. This caused not only 75-80% loss of histaminergic neurons in the posterior hypothalamus, but also some loss of other orexin-receptor expressing cells e.g. MCH neurons. In the long-day anabolic state, lesions produced a transient post-surgical decrease in body weight, but the hamsters recovered and maintained constant body weight, whereas weight gradually increased in sham-lesioned hamsters. VO(2) in the dark phase was significantly higher in the lesioned hamsters compared to shams, and locomotor activity also tended to be higher. In a second study in short days, sham-treated hamsters showed the expected seasonal decrease in body weight, but weight remained constant in the lesioned hamsters, as in the long-day study. Lesioned hamsters consumed more during the early dark phase and less during the light phase due to an increase in the frequency of meals during the dark and decreased meal size during the light, and their cumulative food intake in their home cages was greater than in the control hamsters. In summary, ablation of orexin-responsive cells in the posterior hypothalamus blocks the short-day induced decline in body weight by preventing seasonal hypophagia, evidence consistent with the hypothesis that central histaminergic mechanisms contribute to long-term regulation of body weight.
Puberty is the attainment of fertility -- the ability to reproduce. It occurs because our brains ... more Puberty is the attainment of fertility -- the ability to reproduce. It occurs because our brains begin to secrete one key hormone, gonadotropin-releasing hormone. This causes the pituitary gland to induce maturation of the testis and the ovary, which then produce sex steroid hormones and mature sperm and eggs. We should be concerned that the age at which puberty begins is decreasing in many countries. Puberty starts when adequate growth and energy storage has occurred, so the earlier age of puberty may reflect recent increases in adolescent obesity resulting from more sedentary lifestyles.
bioRxiv (Cold Spring Harbor Laboratory), Dec 16, 2021
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycytespecific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission.
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylasepositive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Singlecell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced expression in the mutant mouse. However, the mild tanycyte depletion and loss of markers observed in NrCAM-deficient mice were associated with only a subtle metabolic phenotype.
Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are ... more Hypothalamic tanycytes are neural stem and progenitor cells, but little is known of how they are regulated. Here we provide evidence that the cell adhesion molecule, NrCAM, regulates tanycytes in the adult niche. NrCAM is strongly expressed in adult mouse tanycytes. Immunohistochemical and in situ hybridization analysis revealed that NrCAM loss of function leads to both a reduced number of tanycytes and reduced expression of tanycyte-specific cell markers, along with a small reduction in tyrosine hydroxylase-positive arcuate neurons. Similar analyses of NrCAM mutants at E16 identify few changes in gene expression or cell composition, indicating that NrCAM regulates tanycytes, rather than early embryonic hypothalamic development. Neurosphere and organotypic assays support the idea that NrCAM governs cellular homeostasis. Single-cell RNA sequencing (scRNA-Seq) shows that tanycyte-specific genes, including a number that are implicated in thyroid hormone metabolism, show reduced express...
Twelve lactating sows were used at 22.4±0.8 days postpartum to determine whether endogenous opioi... more Twelve lactating sows were used at 22.4±0.8 days postpartum to determine whether endogenous opioid peptides (FOP) are involved in the suckling-induced inhibition of luteinizing hormone (LH) secretion. Four sows each received either 1, 2, or 4 mg/kg body weight of naloxone (NA L), an opiate antagonist, in saline i. v. Blood was collected at 15-mm intervals for 2 h before and 4 b after NAL treatment. All sows were then given 100 pg gonadotropin-releasing hormone (GnRH) in saline iv., and blood samples were collected for an additional h. Pigs were weaned after blood sampling. At 40 b after weaning, sows were treated and blood samples collected as during suckling. Serum concentrations of LH after treatment with NAL were similar for all doses; therefore, the data were pooled across doses. During suckling, serum concentrations of LH were 0.41±0.04 ng/ml before NAL treatment, increased to 0.65±0.08 ng/ml at 30 mm after NAL treatment, and remained elevated above pretreatmerit concentrations for 120 mm (p<0.05). Naloxone failed to alter serum concentrations of LH after weaning. Tbese data indicate that EOP may be involved in the suckling-induced suppression of LH secretion and that weaning may either decrease opioid inhibition of LH secretion or decrease pituitary LH responsiveness to endogenous GnRH released by NAL.
4-£ F x X. rt E. S' 3 I? 5' ... Melatonin and the Mammalian Pineal Gland This One 66C3... more 4-£ F x X. rt E. S' 3 I? 5' ... Melatonin and the Mammalian Pineal Gland This One 66C3-RWT-826F ... Melatonin and the Mammalian Pineal Gland Josephine Arendt Professor of Endocrinology School of Biological Sciences University of Surrey Guildford, UK ...
Background and aimTherapeutic activation of thermogenic brown adipose tissue (BAT) is a potential... more Background and aimTherapeutic activation of thermogenic brown adipose tissue (BAT) is a potential strategy to prevent obesity and metabolic disease in humans. However, it is now recognised that rodent studies examining BAT physiology are carried out at sub‐thermoneutral temperatures (e.g. ~20°C), and are not translationally relevant to humans as BAT is ‘hyperactive’. Therefore, the aim of this study was to determine the effect of common regulators of BAT metabolism when animals were raised at thermoneutrality (28°C).Material and methodsThirty weanling Sprague‐Dawley rats were housed at thermoneutrality (28°C) and randomised to either chow (C, n=6) or a high‐fat diet (HFD, n=24) from 3‐weeks of age. At 12 weeks, subgroups (n=6) of HFD were randomised to either mild‐cold exposure (20°C), Mirabegron, a selective β3‐agonist (0.75 mk/kg/d) or exercise training (1h/d, 5 d/week). Metabolic assessment was undertaken in CLAMS during the last 48h to assess energy intake (EI), expenditure (EE) and physical activity (PA) in addition to the acute response to administration of Mirabegron. Key thermogenic and metabolic genes were analysed in interscapular BAT by qPCR in addition to targeted insulin resistance PCR Arrays (86 key genes, n=3).ResultsNo interventions reduced body weight or fat mass. Intriguingly however, mild‐cold exposure significantly increased weight‐gain during the 4‐week period (78.6 vs. 119.8g). This was accompanied by a significant increase in inguinal AT (7.14 vs 16.14g) in cold‐exposed animals whilst BAT mass was significantly increased with exercise‐training (0.74 vs. 1.2g). There was no difference in 24h EE, EI or PA between groups. Key thermogenic genes in BAT were unchanged by the interventions. CITED1 expression was upregulated by HFD and reduced by all interventions whilst PRDM16 expression was reduced by HFD and increased by exercise. Similarly, expression of PPARA, mTOR and the ‘beige’ marker TBX1 were upregulated by exercise only. Targeted PCR arrays demonstrated an upregulation of inflammatory markers e.g. TLR4, EMR1, CASP1 and IL18R1 and a downregulation of metabolic genes e.g. SCD1, FASN, ACACB, HK2 with HFD. β3 increased FASN, whilst IL18R1, IL6 and STAT3 were downregulated along with IRS2, LEP, ADIPOR1, INSR and PIK3R1. Exercise upregulated FASN, SCD1, HK, ACACA, ACACB and PDK2 but NLRP3, IL1β, PYCARD, LPL, IRS2, INSR, FABP4 and CD36 were all downregulated.ConclusionWe show there is no consistent upregulation of BAT as determined by key thermogenic genes in response to common stimuli when examined at thermoneutrality. Effects of interventions on BAT carried out at sub‐thermoneutrality are most likely to be a consequence of chronic mild‐cold stress and are unlikely to be translated to humans.Support or Funding InformationBritish Heart FoundationThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Background and aim: Rodents are commonly housed below thermoneutrality and this exposure to 'cold... more Background and aim: Rodents are commonly housed below thermoneutrality and this exposure to 'cold' (i.e. 20°C) activates thermogenic brown (BAT) and beiging of white adipose tissue. Here, we examined whether a standard housing temperature (i.e. 20°C, a reduction in temperature of ~8°C) or YM-178, a highly-selective β 3-adrenoreceptor agonist, in obese animals raised at thermoneutrality, would impact differently on classical BAT or subcutaneous inguinal (IWAT) beige depots. Methods: Eighteen weanling Sprague-Dawley rats were housed at thermoneutrality (28°C) and fed a high-fat diet. At 12 weeks, 6 animals were randomised to either standard housing temperature (20°C, n=6) or to β 3-AR agonist administration (28°C+β3, 0.75mg/kg/d, n=6) for 4 weeks. Metabolic assessment was undertaken during the final 48h, followed by interscapular, perivascular BAT and IWAT sampling for the analysis of thermogenic genes and the proteome. Results: Exposure to 20°C increased weight gain, BAT and IWAT mass. Proteomic analysis of BAT revealed novel pathways associated with cold-induced weight gain (i.e. histone deacetylation, glycosaminoglycan degradation and glycosphingolipid biosynthesis) whilst β 3adrenoreceptor agonism impacted on proteins involved in skeletal muscle contraction and cell differentiation. IWAT of cold-exposed animals exhibited an enrichment of proteins involved NAD+ binding, plus retinol and tyrosine metabolic pathways whilst β 3-AR agonism downregulated ribosomal and upregulated acute phase response proteins. Conclusion: Following diet-induced obesity at thermoneutrality, exposure to 20°C promotes subcutaneous fat deposition in order to reduce heat loss and defend body temperature. In contrast, chronic administration of β 3-AR agonist has minimal metabolic-related effects on adipose tissue. .
The Siberian hamster (Phodopus sungorus) survives winter by decreasing food intake and catabolizi... more The Siberian hamster (Phodopus sungorus) survives winter by decreasing food intake and catabolizing abdominal fat reserves, resulting in a sustained, profound loss of body weight. Hypothalamic tanycytes are pivotal for this process. In these cells, short-winter photoperiods upregulate deiodinase 3, an enzyme that regulates thyroid hormone availability, and downregulate genes encoding components of retinoic acid (RA) uptake and signaling. The aim of the current studies was to identify mechanisms by which seasonal changes in thyroid hormone and RA signaling from tanycytes might ultimately regulate appetite and energy expenditure. proVGF is one of the most abundant peptides in the mammalian brain, and studies have suggested a role for VGF-derived peptides in the photoperiodic regulation of body weight in the Siberian hamster. In silico studies identified possible thyroid and vitamin D response elements in the VGF promoter. Using the human neuroblastoma SH-SY5Y cell line, we demonstrate...
The viral 2A sequence has become an attractive alternative to the traditional internal ribosomal ... more The viral 2A sequence has become an attractive alternative to the traditional internal ribosomal entry site (IRES) for simultaneous over-expression of two genes and in combination with recombinant adeno-associated viruses (rAAV) has been used to manipulate gene expression in vitro. To develop a rAAV construct in combination with the viral 2A sequence to allow long-term over-expression of the vgf gene and fluorescent marker gene for tracking of the transfected neurones in vivo. Transient transfection of the AAV plasmid containing the vgf gene, viral 2A sequence and eGFP into SH-SY5Y cells resulted in eGFP fluorescence comparable to a commercially available reporter construct. This increase in fluorescent cells was accompanied by an increase in VGF mRNA expression. Infusion of the rAAV vector containing the vgf gene, viral 2A sequence and eGFP resulted in eGFP fluorescence in the hypothalamus of both mice and Siberian hamsters, 32 weeks post infusion. In situ hybridisation confirmed t...
Accurate estimation of the number of ovarian follicles at various stages of development is an imp... more Accurate estimation of the number of ovarian follicles at various stages of development is an important indicator of the process of folliculogenesis in relation to the endocrine signals and paracrine/autocrine mechanisms that control the growth and maturation of the oocytes and their supporting follicular cells. There are 10-fold or greater differences in follicular numbers per ovary at similar ages and/or strains reported in earlier studies using various methods, leading to difficulties with interpretation of ovarian function in control vs experimental conditions. This study describes unbiased, assumption-free stereological methods for quantification of early and growing follicular numbers in the mouse ovary. A fractionator approach was used to sample a defined fraction of histological sections of adult wild-type ovaries. Primordial and primary follicles were counted independently with the optical and physical disector methods. The fractionator/disector methods, which are independe...
We investigated whether histaminergic tone contributes to the seasonal catabolic state in Siberia... more We investigated whether histaminergic tone contributes to the seasonal catabolic state in Siberian hamsters by determining the effect of ablation of histaminergic neurons on food intake, metabolic rate and body weight. A ribosomal toxin (saporin) conjugated to orexin-B was infused into the ventral tuberomammillary region of the hypothalamus, since most histaminergic neurons express orexin receptors. This caused not only 75-80% loss of histaminergic neurons in the posterior hypothalamus, but also some loss of other orexin-receptor expressing cells e.g. MCH neurons. In the long-day anabolic state, lesions produced a transient post-surgical decrease in body weight, but the hamsters recovered and maintained constant body weight, whereas weight gradually increased in sham-lesioned hamsters. VO(2) in the dark phase was significantly higher in the lesioned hamsters compared to shams, and locomotor activity also tended to be higher. In a second study in short days, sham-treated hamsters showed the expected seasonal decrease in body weight, but weight remained constant in the lesioned hamsters, as in the long-day study. Lesioned hamsters consumed more during the early dark phase and less during the light phase due to an increase in the frequency of meals during the dark and decreased meal size during the light, and their cumulative food intake in their home cages was greater than in the control hamsters. In summary, ablation of orexin-responsive cells in the posterior hypothalamus blocks the short-day induced decline in body weight by preventing seasonal hypophagia, evidence consistent with the hypothesis that central histaminergic mechanisms contribute to long-term regulation of body weight.
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