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A13—General Neurobiology-Poster Session
Comparative Biochemistry and Physiology, Part A 143 (2006) S117 – S118 www.elsevier.com/locate/cbpa
Society for Experimental Biology Annual Main Meeting 2nd–7th April 2006, University of Kent at Canterbury, UK
A13–GENERAL NEUROBIOLOGY-POSTER SESSION Organised by P. Newland (University of Southampton)
A13.1 Steroidal natural products from ginseng inhibit sodium channel function by interacting with the alkaloid neurotoxin binding site 2 R. Nicholson, Y. Duan, J. Zheng, V. Law, (Simon Fraser University) Two ginseng aglycones (20(S)protopanaxadiol and 20(S)protopanaxatriol) and a monoglucoside of 20(S)protopanaxadiol (Rh2) were found to displace the binding of the site 2-selective radioligand [3H] batrachotoxinin A-20a-benzoate to sodium channels of mouse brain. Inhibitory potencies were: 20(S)protopanaxadiol (IC50 = 42 AM); 20(S)protopanaxatriol (IC50 = 79 AM) and Rh2 (IC50 = 162 AM). Reversal of inhibition occurred slowly. Scatchard experiments confirmed that 20(S)protopanaxadiol and Rh2 decreased the Kd of radioligand, however Rh2 additionally reduced Bmax. Both compounds increased dissociation of the [3H] batrachotoxinin A-20abenzoate:sodium channel complex, but failed to influence the rate of association of radioligand. Our results indicate that 20(S)protopanaxadiol and Rh2 allosterically destabilize radioligand-activated channels through non-covalent modification of neurotoxin binding site 2. The study compounds also inhibited sodium channel site 2 activatordependent release of neurotransmitters L-glutamate, GABA and aspartate from synaptosomes.
A13.2 Nitric oxide modulates an oviposition digging rhythm of locusts via a PKG-dependent regulatory pathway P. Newland, P. Yates, (University of Southampton) Central pattern generators are not hard-wired circuits, but instead are flexible multifunctional systems capable of producing different motor outputs or can be shaped to suit the demands posed by fluctuating internal and external environments. In this study we have examined the role of nitric oxide (NO) in modulating an oviposition digging rhythm of locusts that is utilised during egg laying. We show that nitric oxide continuously regulates the cycle frequency of the rhythm; increasing the frequency when endogenous and exogenous NO levels doi:10.1016/j.cbpa.2006.01.047
are increased (with NO donors SNAP and PAPANONOate, and Larginine), but decreasing the frequency when NO levels are decreased (using L-NAME and PTIO). We found that NO mediates its effects by acting on one of its molecular targets, the enzyme soluble guanylate cyclase (sGC) to elevate cGMP levels. By using a specific protein kinase G inhibitor (KT5823) we show that the increase in cGMP levels elevates PKG that in turn acts to increase the cycle speed of the digging rhythm. This regulatory mechanism provides a continuous and dynamic control over the digging rhythm that allows it to be shaped to match internal and external conditions. This work was supported by a BBSRC grant to PLN.
A13.3 Nitric oxide adjusts salt responses H. Schuppe, M. Cuttle, P. Newland, (University of Southampton) Locusts require sodium chloride (NaCl) as part of their diet, and detect it with taste receptors on the surface of their body. Previous studies have suggested that one or more of the sensory neurons involved contain soluble guanylate cyclase, one target for nitric oxide (NO). We therefore ask, whether NO contributes to salt taste sensitivity, and whether it plays a role in dietary regulation. In the epidermis of locusts nitric oxide is synthesized by the glandular cells. From there NO may diffuse into taste receptors to exert its effect on sensory neurons. Changing the NO levels within the preparation modulates sensory responses to NaCl. For example, applying drugs that increase NO levels decreases the spike rate of salt responses. Our results suggest that NO is endogenously generated and continuously attenuates the sensitivity of the chemosensory neurons to NaCl. The effect of NO is likely to be mediated by direct binding of NO to sulfhydryl groups, possibly belonging to epithelial sodium channels, and does not involve the NO/cGMP pathway. NO synthesis in glandular cells increases in response to falling salt concentration in the haemolymph. We therefore hypothesize that NO modulates salt sensitivity of taste receptors dependent on haemolymph salt concentration, and suggest that this could affect the uptake of salt and other nutrients. This work was supported by a project grant to P. L. N. from the BBSRC.
S118
Abstracts / Comparative Biochemistry and Physiology, Part A 143 (2006) S117 – S118
A13.4 Does larval diet influence adult food choice? Gustatory learning and memory in Drosophila melanogaster S. Young, P. Newland, D. Shepherd, H. Schuppe, G. Poppy, (University of Southampton) Gustation underlies much insect behaviour, including finding and assessing food and egg laying sites. Drosophila are capable of learning sensory information about their environment, however, whether they are capable of learning gustatory information and using this information to modify their adult behaviour is as yet unknown. A behavioural assay was therefore designed to try to test this issue. Drosophila larvae were raised on food media with carbohydrate concentration and type varied. Individuals were removed from these diets as pupae and as adults given the choice to feed on either the diet upon which they were raised or a standard diet. Results show that adult individuals raised as larvae on food media lacking sucrose have a preference to feed on this medium over a medium containing sucrose. When raised as larvae on other concentrations of sucrose, however, adult preference is always for the highest carbohydrate content medium. Nevertheless adults are more likely to accept the reduced carbohydrate content medium if they were exposed to it as larvae. Adult individuals raised as larvae on reduced concentrations of trehalose were less accepting of these media as adults compared to those raised as larvae on a standard trehalose diet. Varying the concentration of fructose in the larval diet had no subsequent effect on adult feeding preferences. In summary, results show that varying larval diet can affect subsequent adult feeding preferences, although the effects depend on the constituent of the diet that is being varied. Possible reasons for these modifications in behaviour are discussed.
A13.5 Stress induced reduction of neurogenesis in rainbow trout brain C. Sørensen, G. Nilsson, (University of Oslo); Ø. Øverli, (Norwegian University of Life Sciences) Adult neurogenesis, the production of new neurons in the cns of adult animals, has been demonstrated in all examined vertebrates. In mammals this process appears to be limited to a few discrete brain areas, most notably the hippocampus. Over the last decade it has become clear that the rate of hippocampal neurogenesis is affected by several factors, including environmental complexity, physical exercise, learning, and hormone levels. The rate of neurogenesis does in turn appear to influence the hippocampal function in memory formation and mood regulation. In several animal models, chronic stress has been associated with reduced neurogenesis, and reduced
hippocampal volumes have been found in humans with severe depressive disorders. In non-mammalian vertebrates the anatomical aspects of adult neurogenesis has been investigated, but few studies have focused on the functional aspects of adult neurogenesis. In this study, a wellcharacterized model system for social stress in rainbow trout (Oncorhynchus mykiss) was used to investigate the effect of chronic stress on brain cell proliferation. Juvenile rainbow trout were allowed to form social hierarchies, where subordinate animals were subjected to severe and chronic stress. Cell proliferation in the telencephalon, the proposed site of hippocampal function in teleosts, was quantified using BrdU, and a 40% reduction in proliferative rate was found in subordinates compared to unstressed controls. This implies that adult neurogenesis is regulated in teleosts as well as in mammals, and that regulation of hippocampal neurogenic rate might be a common means of modifying hippocampal function in all vertebrates.
A13.6 Neuropeptide-containing neurons in the central nervous system of the waterflea Daphnia magna H. Dircksen, (Stockholm University); Q. Zhang, (Anatomical Institute Frankfurt University) Neuronal structures immunoreactive to antisera against neuropeptides of decapod crustaceans were immunostained in whole mounts and sections of the central nervous system (CNS) of Daphnia magna. Classical decapod-like X-organ-sinus gland structures known to contain crustacean hyperglycemic hormone (CHH), red pigmentconcentrating hormone (RPCH), and pigment-dispersing hormone (PDH) were not found in Daphnia but only peripheral putative neurohaemal areas (NHA) in the thorax and/or distinct CNS interneurons for these peptides. The only possible exception is a putative NHA formed at the ventral brain by prominently staining locust adipokinetic hormone-immunoreactive neurons. Two pairs of CHH-IR interneurons are detected in the brain; others occur in the peripheral nervous system of the ventral body at the base of the legs. Five bipolar RPCH-IR interneurons only occur in the thoracic ganglia with extensions into the suboesophageal ganglion and the brain. PDHIR neurons are confined to the brain and the optic ganglia. Two CCAP-IR neurones only located in the subsesophageal ganglion ascend to the posterior brain and descend to the caudal end of the CNS, and terminate at the tail spines. Biochemical evidence (HPLC, ELISA) for the existence of true decapod-like neuropeptides substantiated by negative staining reactions observed in preabsorption controls led us to assume that the neuropeptides in Cladocera are structurally closely related to those of malacostracan orders but may have partially divergent functions.
Society for Experimental Biology Annual Main Meeting 2nd–7th April 2006, University of Kent at Canterbury, UK
A13–GENERAL NEUROBIOLOGY-POSTER SESSION Organised by P. Newland (University of Southampton)
A13.1 Steroidal natural products from ginseng inhibit sodium channel function by interacting with the alkaloid neurotoxin binding site 2 R. Nicholson, Y. Duan, J. Zheng, V. Law, (Simon Fraser University) Two ginseng aglycones (20(S)protopanaxadiol and 20(S)protopanaxatriol) and a monoglucoside of 20(S)protopanaxadiol (Rh2) were found to displace the binding of the site 2-selective radioligand [3H] batrachotoxinin A-20a-benzoate to sodium channels of mouse brain. Inhibitory potencies were: 20(S)protopanaxadiol (IC50 = 42 AM); 20(S)protopanaxatriol (IC50 = 79 AM) and Rh2 (IC50 = 162 AM). Reversal of inhibition occurred slowly. Scatchard experiments confirmed that 20(S)protopanaxadiol and Rh2 decreased the Kd of radioligand, however Rh2 additionally reduced Bmax. Both compounds increased dissociation of the [3H] batrachotoxinin A-20abenzoate:sodium channel complex, but failed to influence the rate of association of radioligand. Our results indicate that 20(S)protopanaxadiol and Rh2 allosterically destabilize radioligand-activated channels through non-covalent modification of neurotoxin binding site 2. The study compounds also inhibited sodium channel site 2 activatordependent release of neurotransmitters L-glutamate, GABA and aspartate from synaptosomes.
A13.2 Nitric oxide modulates an oviposition digging rhythm of locusts via a PKG-dependent regulatory pathway P. Newland, P. Yates, (University of Southampton) Central pattern generators are not hard-wired circuits, but instead are flexible multifunctional systems capable of producing different motor outputs or can be shaped to suit the demands posed by fluctuating internal and external environments. In this study we have examined the role of nitric oxide (NO) in modulating an oviposition digging rhythm of locusts that is utilised during egg laying. We show that nitric oxide continuously regulates the cycle frequency of the rhythm; increasing the frequency when endogenous and exogenous NO levels doi:10.1016/j.cbpa.2006.01.047
are increased (with NO donors SNAP and PAPANONOate, and Larginine), but decreasing the frequency when NO levels are decreased (using L-NAME and PTIO). We found that NO mediates its effects by acting on one of its molecular targets, the enzyme soluble guanylate cyclase (sGC) to elevate cGMP levels. By using a specific protein kinase G inhibitor (KT5823) we show that the increase in cGMP levels elevates PKG that in turn acts to increase the cycle speed of the digging rhythm. This regulatory mechanism provides a continuous and dynamic control over the digging rhythm that allows it to be shaped to match internal and external conditions. This work was supported by a BBSRC grant to PLN.
A13.3 Nitric oxide adjusts salt responses H. Schuppe, M. Cuttle, P. Newland, (University of Southampton) Locusts require sodium chloride (NaCl) as part of their diet, and detect it with taste receptors on the surface of their body. Previous studies have suggested that one or more of the sensory neurons involved contain soluble guanylate cyclase, one target for nitric oxide (NO). We therefore ask, whether NO contributes to salt taste sensitivity, and whether it plays a role in dietary regulation. In the epidermis of locusts nitric oxide is synthesized by the glandular cells. From there NO may diffuse into taste receptors to exert its effect on sensory neurons. Changing the NO levels within the preparation modulates sensory responses to NaCl. For example, applying drugs that increase NO levels decreases the spike rate of salt responses. Our results suggest that NO is endogenously generated and continuously attenuates the sensitivity of the chemosensory neurons to NaCl. The effect of NO is likely to be mediated by direct binding of NO to sulfhydryl groups, possibly belonging to epithelial sodium channels, and does not involve the NO/cGMP pathway. NO synthesis in glandular cells increases in response to falling salt concentration in the haemolymph. We therefore hypothesize that NO modulates salt sensitivity of taste receptors dependent on haemolymph salt concentration, and suggest that this could affect the uptake of salt and other nutrients. This work was supported by a project grant to P. L. N. from the BBSRC.
S118
Abstracts / Comparative Biochemistry and Physiology, Part A 143 (2006) S117 – S118
A13.4 Does larval diet influence adult food choice? Gustatory learning and memory in Drosophila melanogaster S. Young, P. Newland, D. Shepherd, H. Schuppe, G. Poppy, (University of Southampton) Gustation underlies much insect behaviour, including finding and assessing food and egg laying sites. Drosophila are capable of learning sensory information about their environment, however, whether they are capable of learning gustatory information and using this information to modify their adult behaviour is as yet unknown. A behavioural assay was therefore designed to try to test this issue. Drosophila larvae were raised on food media with carbohydrate concentration and type varied. Individuals were removed from these diets as pupae and as adults given the choice to feed on either the diet upon which they were raised or a standard diet. Results show that adult individuals raised as larvae on food media lacking sucrose have a preference to feed on this medium over a medium containing sucrose. When raised as larvae on other concentrations of sucrose, however, adult preference is always for the highest carbohydrate content medium. Nevertheless adults are more likely to accept the reduced carbohydrate content medium if they were exposed to it as larvae. Adult individuals raised as larvae on reduced concentrations of trehalose were less accepting of these media as adults compared to those raised as larvae on a standard trehalose diet. Varying the concentration of fructose in the larval diet had no subsequent effect on adult feeding preferences. In summary, results show that varying larval diet can affect subsequent adult feeding preferences, although the effects depend on the constituent of the diet that is being varied. Possible reasons for these modifications in behaviour are discussed.
A13.5 Stress induced reduction of neurogenesis in rainbow trout brain C. Sørensen, G. Nilsson, (University of Oslo); Ø. Øverli, (Norwegian University of Life Sciences) Adult neurogenesis, the production of new neurons in the cns of adult animals, has been demonstrated in all examined vertebrates. In mammals this process appears to be limited to a few discrete brain areas, most notably the hippocampus. Over the last decade it has become clear that the rate of hippocampal neurogenesis is affected by several factors, including environmental complexity, physical exercise, learning, and hormone levels. The rate of neurogenesis does in turn appear to influence the hippocampal function in memory formation and mood regulation. In several animal models, chronic stress has been associated with reduced neurogenesis, and reduced
hippocampal volumes have been found in humans with severe depressive disorders. In non-mammalian vertebrates the anatomical aspects of adult neurogenesis has been investigated, but few studies have focused on the functional aspects of adult neurogenesis. In this study, a wellcharacterized model system for social stress in rainbow trout (Oncorhynchus mykiss) was used to investigate the effect of chronic stress on brain cell proliferation. Juvenile rainbow trout were allowed to form social hierarchies, where subordinate animals were subjected to severe and chronic stress. Cell proliferation in the telencephalon, the proposed site of hippocampal function in teleosts, was quantified using BrdU, and a 40% reduction in proliferative rate was found in subordinates compared to unstressed controls. This implies that adult neurogenesis is regulated in teleosts as well as in mammals, and that regulation of hippocampal neurogenic rate might be a common means of modifying hippocampal function in all vertebrates.
A13.6 Neuropeptide-containing neurons in the central nervous system of the waterflea Daphnia magna H. Dircksen, (Stockholm University); Q. Zhang, (Anatomical Institute Frankfurt University) Neuronal structures immunoreactive to antisera against neuropeptides of decapod crustaceans were immunostained in whole mounts and sections of the central nervous system (CNS) of Daphnia magna. Classical decapod-like X-organ-sinus gland structures known to contain crustacean hyperglycemic hormone (CHH), red pigmentconcentrating hormone (RPCH), and pigment-dispersing hormone (PDH) were not found in Daphnia but only peripheral putative neurohaemal areas (NHA) in the thorax and/or distinct CNS interneurons for these peptides. The only possible exception is a putative NHA formed at the ventral brain by prominently staining locust adipokinetic hormone-immunoreactive neurons. Two pairs of CHH-IR interneurons are detected in the brain; others occur in the peripheral nervous system of the ventral body at the base of the legs. Five bipolar RPCH-IR interneurons only occur in the thoracic ganglia with extensions into the suboesophageal ganglion and the brain. PDHIR neurons are confined to the brain and the optic ganglia. Two CCAP-IR neurones only located in the subsesophageal ganglion ascend to the posterior brain and descend to the caudal end of the CNS, and terminate at the tail spines. Biochemical evidence (HPLC, ELISA) for the existence of true decapod-like neuropeptides substantiated by negative staining reactions observed in preabsorption controls led us to assume that the neuropeptides in Cladocera are structurally closely related to those of malacostracan orders but may have partially divergent functions.