- Email: [email protected]
HIGHLIGHTS IN BASIC AUTONOMIC NEUROSCIENCES
Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
Contents lists available at ScienceDirect
Autonomic Neuroscience: Basic and Clinical j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / a u t n e u
HIGHLIGHTS IN BASIC AUTONOMIC NEUROSCIENCES Prepared by: James J. Galligan, James A. Brock Section Editor: Michael Gilbey
Sawada, K., Echigo, N., Juge, N., Miyaji, T., Otsuka, M., Omote, H., Yamamoto, A., Moriyama, Y. (Nagahama, Japan). 2008. Identification of a vesicular nucleotide transporter. Proc Natl Acad Sci USA 105, 5683–5686. Article summary There is substantial functional evidence that ATP is a neurotransmitter released by some neurons in the central and peripheral nervous system. However, because ATP is a ubiquitous molecule that is contained in all neurons, it has not been possible to identify a reliable and selective neurochemical marker for purinergic neurons. Sawada and co-workers may have helped solve this problem through the identification of a vesicular nucleotide transporter. This transporter belongs to the family of solute carrier proteins and is encoded by the SLC17A9 gene. This gene encodes an anion transporter and ATP appears to be a substrate for this transporter. Northern blot analysis revealed that the mRNA encoding the proposed nucleotide transporter was found in the brain, adrenal gland, thyroid, liver and lung. Immunocytochemical studies revealed the presence of the transporter in mouse and bovine adrenal gland and immunoreactivity for the transporter was associated with granules in the adrenal chromaffin cells. When the SLC17A9 protein was inserted into proteoliposomes, it transported ATP in a membrane potential dependent manner. Finally, the authors used pheochromacytoma (PC)-12 cells as a model for adrenal chromaffin cells. siRNA knockdown caused a 50% decrease in SLC17A9 expression and a 50% reduction in high K+ stimulated ATP release from treated PC-12 cells. Taken together these data indicate the SLC17A9 protein is a nucleotide transporter that may be responsible for accumulating ATP and other nucleotides in synaptic vesicles and chromaffin granules. Commentary ATP is a neurotransmitter released by peripheral autonomic nerves. It is released with norepinephrine by sympathetic nerves supplying many blood vessels and the vas deferens where ATP causes smooth muscle contraction. ATP is also released with acetylcholine by parasympathetic nerves supplying the bladder where ATP causes bladder contractions. Finally ATP is a fast excitatory transmitter in the enteric nervous system where it is a co-transmitter with acetylcholine at some synapses and it is also an inhibitory neurotransmitter released by nerves supplying the muscle layers. There are numerous neurochemical markers for noradrenergic, cholinergic and neuropeptide containing autonomic nerves but the relationship of these nerves 1566-0702/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.autneu.2009.07.012
to purinergic nerves has not been established using neuroanatomical techniques. Functional data indicate that ATP is co-released with norepinephrine or acetylcholine but it is not clear if these neurotransmitters are co-stored in the same synaptic vesicles or the same nerve terminals in all cases. The SLC17A9 protein can be used to generate antibodies against this putative nucleotide transporter and this raises the possibility that immunocytochemical approaches can be used to identify purinergic nerves. Co-staining with markers for the vesicular monoamine and vesicular acetylcholine transporters will allow determination of whether monoamine or acetylcholine containing vesicles also store ATP. It may also be possible to develop blockers of the vesicular nucleotide transporter (similar to vesamicol for the vesicular acetylcholine transporter or reserpine for the monoamine transporter) that will allow studies of the effects of selective depletion of vesicular ATP stores on autonomic neurotransmission. It remains to be determined if the SLC17A9 protein functions as a nucleotide transporter in autonomic nerves as it appears to do in adrenal chromaffin cells. However, in the event that the SLC17A9 is not a neuronal vesicular nucleotide transporter, the data of Sawada et al. may help to narrow the search to structurally similar proteins which would facilitate a successful search for the neuronal nucleotide transporter.
Usui, D., Yamaguchi-Shima, N., Okada, S., Shimizu, T., Wakiguchi, H., Yokotani, K. (Nankoku, Kochi, Japan). 2009. Selective activation of the sympathetic ganglia by centrally administered corticotropin-releasing factor in rats. Auton Neurosci 146, 111–114. Article summary Central nervous system corticotrophin releasing factor (CRF) pathways participate in the stress response in part through their influence on the activity of the autonomic nervous system. In this study, Usui and colleagues administered CRF into the cerebral ventricles (i.c.v.) of anesthetized rats. Thirty and 60 min after CRF administration, the stellate, superior cervical and celiac ganglia were collected from euthanized animals and the ganglia were prepared for immunocytochemical localization of dopamine-β-hydroxylase (DβH; a marker for postganglionic sympathetic neurons) and c-Fos protein (a marker for activated neurons). The authors found that CRF treatment increased c-Fos immunoreactivity in the nuclei of neurons in the stellate and celiac ganglia but not in the superior cervical ganglion. The stimulatory effect was most prominent in the celiac
2
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
ganglion. These data show that central CRF does not produce a generalized autonomic activation but instead produces a selective activation of sympathetic preganglionic neurons that supply specific sympathetic ganglia and perhaps organ specific subsets of neurons within those activated ganglia. Commentary Stress produces a wide variety of responses that involve multiple organ systems. The autonomic nervous system is a major player in the stress response and there are many autonomic manifestations of stress. These include changes in cardiovascular, respiratory tract, gastrointestinal tract and urinary tract functions. CRF is released from hypothalamic neurons into the hypophyseal portal blood supply of the pituitary gland where it stimulates release of adrenocorticotropin which then stimulates release of cortisol from the adrenal cortex. However, increased activity of CRF neurons controlling central autonomic outflow is also part of the stress response. Usui et al. show that centrally administered CRF selectively activates subsets of sympathetic preganglionic neurons. This results in activation of neurons in the stellate ganglion (which supply the heart and lungs) and the celiac ganglion (which supply splanchnic organs) but not the superior cervical ganglion (which largely supplies targets in the head and neck region). These data indicate that central CRF does not produce an overall autonomic arousal but instead there is some selectivity in the pathways activated by CRF. It is also interesting that central CRF did not activate all neurons within the stellate or celiac ganglia. It is not likely that insufficient CRF was administered to cause maximum activation as the authors used 1.5 and 3.0 nM doses. The high dose did not produce a greater neuronal activation than the lower dose. Also, there was no difference in the number of neurons activated at 60 and 120 min post CRF administration, indicating that sufficient time was allowed for maximum activation of the CRF sensitive pathways. These data indicate that there are specific sympathetic postganglionic neurons targeted by CRF sensitive preganglionic pathways. It will be important to identify the specific subsets of neurons activated by CRF as this will provide new details about the organization of the autonomic nervous system and its specific role in the stress response.
Metzger, M., Bareiss, P.M., Danker, T., Wagner, S., Hennenlotter, J., Guenther, E., Obermayr, F., Stenzl, A., Koenigsrainer, A., Skutella, T., Just, L. (Tuebingen, Germany). 2009. Expansion and differentiation of neural progenitors derived from the human adult enteric nervous system. Gastroenterology. Article summary There are a number of gastrointestinal motility and secretory disorders that result from congenital defects in the development and maturation of the enteric nervous system. These defects include Hirschprung's disease and chronic intestinal idiopathic pseudo-obstruction. Surgery is the only treatment and this does not always result in satisfactory outcomes. There is a need for the development of other treatments and transplantation of neural stem cells holds some promise. The work by Metzger and colleagues shows that it may be possible to isolate neural stem and progenitor cells from the enteric nervous system of the adult gastrointestinal tract and these isolated cells could potentially be used in cell-based therapies for developmental abnormalities in the enteric nervous system. Human enteric spheroids were generated from explants isolated from surgical specimens of the small and large intestine that had been removed from adult patients (26– 84 years old) undergoing surgery for cancer, diverticulitis and bladder reconstruction. The authors found that the enterospheroids could be
expanded in culture and spheroid-derived cells could be differentiated into neuronal subtypes after incubation with conditioned medium obtained from murine embryonic gut cultures. These neuronal subtypes included nitrergic, cholinergic and serotonergic neurons. Glial cells could also be identified. Whole cell voltage-clamp studies revealed that differentiated neurons expressed functional voltage-gated sodium and potassium channels. In some experiments, enterospheres were grafted to hindgut segments obtained from E11 day mouse embryos Enteric neurons have not populated the hind gut at this time. Individual cells migrated from the grafted enterosphere into aganglionic explants of mouse intestine and they expressed neuronal markers and had neurone-like extensions. These data indicate that it may be possible to harvest multipotent progenitor cells from the human intestine and that these cells can differentiate into subtypes of enteric neurons or glia. It may be possible to use these enterospheres in cell-based therapies of gastrointestinal motility and secretory disorders that are due to defects in the maturation of the enteric nervous system. Commentary Chronic gastrointestinal motility disorders are very common and unfortunately there are few effective drug treatments for these problems. In most of the patients suffering from these problems, there is no obvious defect in the nerve supply of the gut. However, there are a smaller number of patients who suffer from motility, secretory and absorptive disorders that are based on clear deficits in the nerve supply of the gut. The best known of these is Hirschprung's disease in which the developing enteric nervous system fails to populate the distal colon. Chronic intestinal pseudo-obstruction (CIPO) is a rare but extremely debilitating motility disorder that appears to be due to deficits in the enteric nervous system supplying portions of the small intestine. There are currently no effective treatments of CIPO other than to resect the paralyzed portion of the intestine. The work of Metzger and colleagues may provide some hope for these patients. They have demonstrated the feasibility of isolating and growing neural/glial progenitor cells from adult intestine and that these can be induced to differentiate into subtypes of enteric neurons or glial cells under appropriate conditions. The great advantage of their approach is that specimens of adult human intestine are the source material of which there is a ready supply. Resection of human intestine is performed for a variety of reasons and these tissues are readily available at most academic medical centers. It remains to be seen if these progenitor cells can be induced to differentiate into the full range of neuronal phenotypes present within the intestinal wall and whether they perform the functions of these neurons in re-populated tissues. Can they form synapses with other neurons? Can they maintain appropriate polarity of projection or find appropriate targets (muscle, other neurons, blood vessels, epithelial cells). There are many remaining issues but this study provides important new information about a potential treatment strategy for developmental defects of the enteric nervous system.
Nogueiras, R., Pérez-Tilve, D., Veyrat-Durebex, C., Morgan, D.A., Varela, L., Haynes, W.G., Patterson, J.T., Disse, E., Pfluger, P.T., López, M., Woods, S.C., DiMarchi, R., Diéguez, C., Rahmouni, K., Rohner-Jeanrenaud, F., Tschöp, M.H. (Cincinnati, OH USA). 2009. Direct control of peripheral lipid deposition by CNS GLP-1 receptor signaling is mediated by the sympathetic nervous system and blunted in diet-induced obesity. J Neurosci 29, 5916–5925. Article summary Glucagon-like peptide 1 (GLP-1) is a gut hormone released in response to nutrient absorption. GLP-1 is released into the circulation and it can cross the blood brain barrier. It may act on central nerve circuits involved in energy balance. GLP-1 is also found in neurons in the
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
brain that are involved in energy balance and food intake. The work of Nogueiras et al. investigated the contribution of central GLP-1 receptors in brain pathways regulating fat metabolism by adipocytes independent of feeding behavior. These investigators used lean and obese mice and mice lacking β-adrenergic receptors. They measured sympathetic nerve activity and the expression of enzymes involved in lipid metabolism. In wild-type mice, intracerebroventricular (i.c.v.) infusion of a GLP-1 analog decreased lipid storage in white adipose tissue and this effect was independent from food intake. This effect of i.c.v. GLP-1 was reduced in obese mice and in mice with β1, β2 and β3 adrenergic receptors knocked out. β-receptors (in particular β1 and β2) are known to mediate the regulation of adipocyte lipid metabolism by the sympathetic nervous system. Recordings of the activity of sympathetic nerves supplying white adipose tissue (epididymal fat pad) showed that i.c.v. GLP-1 produced a concentration dependent increase in sympathetic nerve activity. Therefore, the CNS GLP-1 control of adipocyte lipid metabolism appears to be mediated, at least in part, by the sympathetic nervous system and is independent of parallel changes in food intake and body weight. Importantly, the CNS GLP-1 system loses the capacity to modulate adipocyte metabolism in obese states, suggesting an obesity-induced adipocyte resistance to CNS GLP-1. Commentary Obesity is a major public health issue that is likely to become more serious in the future. While poor lifestyle decisions are a major contributor to obesity, there are physiological contributions as well. Understanding the physiological processes that control appetite, fat deposition and energy utilization can only help to understand the causes of obesity and also help to develop preventative treatment strategies. It is well established that activation of the sympathetic nervous system can mobilize fat stores and increase circulating free fatty acids however not all of the mechanisms responsible for sympathetic activation under different physiological conditions are known. GLP-1 is a hormone and central neurotransmitter involved in regulation of energy homeostasis. GLP-1 also regulates appetite. This paper demonstrates that GLP-1 is linked to central sympathetic activation resulting in fat mobilization from white adipose tissue. This response is mediated by β1 and β2 adrenergic receptors expressed by adipocytes. Perhaps the most important finding is that GLP-1 induced activation of the sympathetic nervous system and fat mobilization is impaired in diet-induced obesity at least in mice. Restoration of this mechanism in obese individuals might be a useful approach to the treatment of diet-induced obesity.
Ajmo Jr., C.T., Collier, L., Leonardo, C.C., Hall, A.A., Green, S.M., Womble, T.A., Cuevas, J., Alison, A.E., Pennypacker, K.R. (Tampa, Fl, USA). 2009. Blockade of adrenoreceptors inhibits the splenic response to stroke. Exp Neurol 2009, 218: 47–55. Article summary Previously these authors have demonstrated, in the permanent middle cerebral artery occlusion (MCAO) model of stroke, that splenectomy 2 weeks prior to MCAO reduced the size of the infarction by N80%, highlighting the importance of this immune organ in the generation of secondary injury. They have also demonstrated that MCAO is followed by a reduction in spleen size and that this is associated with an increase in circulating regulatory T cells and macrophages. This study examines whether circulating or nerve-released catecholamines regulate the splenic response to stroke. Rats either underwent splenic denervation 2 weeks prior to MCAO or received α-adrenoceptor antagonists carvedilol (blocks α- and β-receptors), prazosin (α1-receptors) or propranolol (β-receptors). Denervation (confirmed by a marked reduction in TH labeled axons) prior to MCAO did not alter infarct volume or
3
spleen size. Propranolol treatment also had no effects on these outcomes. Prazosin or carvedilol treatment prevented the reduction in spleen size but only carvedilol significantly reduced infarct volume. The authors conclude that blood borne catecholamines released from the adrenal medulla regulate the splenic response to stroke through the activation of both α- and β-receptors. Commentary A prominent inflammatory response occurs following both ischemic and hemorrhagic stroke, thereby exacerbating the injury. There is also growing evidence that systemic inflammation influences stroke outcome and that therapies may need to also attenuate systemic inflammation to be effective. Indeed, the beneficial effects of stem cell therapy may be mediated, at least in part, by its systemic antiinflammatory effects. For example the authors have previously demonstrated that systemic treatment with human umbilical cord blood cells after MCAO prevents the reduction in spleen size, promotes expression of anti-inflammatory cytokines in the spleen and reduces neutrophil infiltration at the infarct site in the MCAO model. In this paper, the authors have investigated the involvement of the sympathetic nervous system in triggering the response of the spleen to MCAO. The findings demonstrate that carvedilol, possibly through a combined blockade of α- and β-adrenoceptors, reduces secondary injury at the infarct site. As blockade of α-adrenoceptors alone did not limit infarct volume but prevented the stroke-induced reduction in spleen size, these two events can be dissociated. Instead the authors suggest a role for circulating catecholamines in mediating the pro-inflammatory effects produced by MCAO in the spleen but do not identify their site(s) of action. This contrasts with the known anti-inflammatory action of activating β2adrenoceptors expressed on innate and adaptive immune cells; activation of these receptors by nerve-released and circulating catecholamines is believed to contribute to post-stroke immune suppression. The importance of this paper is that it demonstrates that stroke therapies should not focus solely on the central nervous system, but also target the peripheral immune cell signaling that plays a key role in ischemic injury progression. This will require greater understanding of role the sympathetic nervous system plays in regulating immune function.
Bove, G.M. (Portland, ME, USA). Focal nerve inflammation induces neuronal signs consistent with symptoms of early complex regional pain syndromes. Exp Neurol (Epub, ahead of print, 2009). Article summary During the early (acute) phase of complex regional pain syndromes (CRPS) there is both severe pain and autonomic dysfunction in the affected limb. While CRPS can be caused by overt nerve injury (type II), it can also result from relatively minor trauma (type I). Inflammation has been implicated in the etiology of CRPS. The author hypothesizes that inflammation of a nerve proximal to the affected region may result in changes that are consistent with clinical CRPS. To investigate this possibility he used a rat model of neuritis (induced using Freund's complete adjuvant) and recorded the activity of single cutaneous sensory and sympathetic axons at a point distal to the site of inflammation (in sural nerve following induction of neuritis in the sciatic nerve). In control rats, no sensory axons had ongoing activity and mean frequency of action potential discharge in sympathetic axons was 2.26 ± 1.33 Hz (mean ± SD). In contrast, in rats with inflamed nerves, 27% of slowly conducting sensory axons (b4 m/s: including Aδ- and C-neurons) had ongoing activity after 3–4 days, and 50% had such activity after 7–8 days. The sympathetic axons neurons had lower frequencies of ongoing discharge following induction of
4
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
neuritis (1.96 ± 1.19 Hz after 3–4 days, 1.48 ± 1.23 Hz after 7–8 days). These findings demonstrate that focal nerve inflammation of a nerve trunk is sufficient to cause changes in the activity of both sensory and sympathetic axons projecting to skin. Commentary The clinical features of CRPS are pain, impaired motor function, edema and abnormalities due to altered sympathetic nerve activity (changes in sweating and blood flow). Furthermore, in some cases, ‘coupling’ between the sympathetic and sensory nervous systems plays a role in maintaining ongoing pain, although the precise locus for this interaction is not known. There is growing evidence that the initial insult results in changes to the central nervous system that modify sensory, motor and autonomic function. In patients with acute CRPS I, Baron and colleagues provided evidence that activity of sympathetic vasoconstrictor neurons is unilaterally reduced, causing swelling and redness of the affected limb (see the papers Discussion). These investigators have also presented evidence that sympathetic nerve activity is reduced during chronic CRPS I, although during this phase skin temperature and blood flow are reduced indicating
considerable cutaneous vasoconstriction. This paradox may be explained by ‘supersensitivity’ of the vasculature induced by the lower levels of sympathetic nerve activation (like decentralizationinduced supersensitivity). Furthermore, reduced levels of ongoing sympathetic nerve activity have been implicated in the acquisition of adrenergic sensitivity by nociceptors. In this paper the investigator has for the first time investigated the effects of experimental neuritis on sympathetic nerve activity. In previous papers, he has demonstrated that focal inflammation with Freund's complete adjuvant induces axonal mechano-sensitivity of intact deep somatic afferent A- and C-fibers and that this ‘sensitization’ is almost absent in cutaneous afferents. However, the present study indicates that focal neuritis induces ectopic activity in many of the C- and slowly conducting Aδ-afferent fibers supplying skin of the rat hindpaw (up to 50%; likely to be mostly nociceptive). As has been suggested in CRPS, the centrally determined level of ongoing activity in cutaneous sympathetic vasoconstrictor axons (baroreceptor sensitive) was concomitantly reduced. These changes would be expected to cause pain and contribute to hyperemia and swelling, perceived and expressed in the territory supplied by these axons (i.e. the hindpaw) and this now needs to be investigated.
Contents lists available at ScienceDirect
Autonomic Neuroscience: Basic and Clinical j o u r n a l h o m e p a g e : w w w. e l s ev i e r. c o m / l o c a t e / a u t n e u
HIGHLIGHTS IN BASIC AUTONOMIC NEUROSCIENCES Prepared by: James J. Galligan, James A. Brock Section Editor: Michael Gilbey
Sawada, K., Echigo, N., Juge, N., Miyaji, T., Otsuka, M., Omote, H., Yamamoto, A., Moriyama, Y. (Nagahama, Japan). 2008. Identification of a vesicular nucleotide transporter. Proc Natl Acad Sci USA 105, 5683–5686. Article summary There is substantial functional evidence that ATP is a neurotransmitter released by some neurons in the central and peripheral nervous system. However, because ATP is a ubiquitous molecule that is contained in all neurons, it has not been possible to identify a reliable and selective neurochemical marker for purinergic neurons. Sawada and co-workers may have helped solve this problem through the identification of a vesicular nucleotide transporter. This transporter belongs to the family of solute carrier proteins and is encoded by the SLC17A9 gene. This gene encodes an anion transporter and ATP appears to be a substrate for this transporter. Northern blot analysis revealed that the mRNA encoding the proposed nucleotide transporter was found in the brain, adrenal gland, thyroid, liver and lung. Immunocytochemical studies revealed the presence of the transporter in mouse and bovine adrenal gland and immunoreactivity for the transporter was associated with granules in the adrenal chromaffin cells. When the SLC17A9 protein was inserted into proteoliposomes, it transported ATP in a membrane potential dependent manner. Finally, the authors used pheochromacytoma (PC)-12 cells as a model for adrenal chromaffin cells. siRNA knockdown caused a 50% decrease in SLC17A9 expression and a 50% reduction in high K+ stimulated ATP release from treated PC-12 cells. Taken together these data indicate the SLC17A9 protein is a nucleotide transporter that may be responsible for accumulating ATP and other nucleotides in synaptic vesicles and chromaffin granules. Commentary ATP is a neurotransmitter released by peripheral autonomic nerves. It is released with norepinephrine by sympathetic nerves supplying many blood vessels and the vas deferens where ATP causes smooth muscle contraction. ATP is also released with acetylcholine by parasympathetic nerves supplying the bladder where ATP causes bladder contractions. Finally ATP is a fast excitatory transmitter in the enteric nervous system where it is a co-transmitter with acetylcholine at some synapses and it is also an inhibitory neurotransmitter released by nerves supplying the muscle layers. There are numerous neurochemical markers for noradrenergic, cholinergic and neuropeptide containing autonomic nerves but the relationship of these nerves 1566-0702/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.autneu.2009.07.012
to purinergic nerves has not been established using neuroanatomical techniques. Functional data indicate that ATP is co-released with norepinephrine or acetylcholine but it is not clear if these neurotransmitters are co-stored in the same synaptic vesicles or the same nerve terminals in all cases. The SLC17A9 protein can be used to generate antibodies against this putative nucleotide transporter and this raises the possibility that immunocytochemical approaches can be used to identify purinergic nerves. Co-staining with markers for the vesicular monoamine and vesicular acetylcholine transporters will allow determination of whether monoamine or acetylcholine containing vesicles also store ATP. It may also be possible to develop blockers of the vesicular nucleotide transporter (similar to vesamicol for the vesicular acetylcholine transporter or reserpine for the monoamine transporter) that will allow studies of the effects of selective depletion of vesicular ATP stores on autonomic neurotransmission. It remains to be determined if the SLC17A9 protein functions as a nucleotide transporter in autonomic nerves as it appears to do in adrenal chromaffin cells. However, in the event that the SLC17A9 is not a neuronal vesicular nucleotide transporter, the data of Sawada et al. may help to narrow the search to structurally similar proteins which would facilitate a successful search for the neuronal nucleotide transporter.
Usui, D., Yamaguchi-Shima, N., Okada, S., Shimizu, T., Wakiguchi, H., Yokotani, K. (Nankoku, Kochi, Japan). 2009. Selective activation of the sympathetic ganglia by centrally administered corticotropin-releasing factor in rats. Auton Neurosci 146, 111–114. Article summary Central nervous system corticotrophin releasing factor (CRF) pathways participate in the stress response in part through their influence on the activity of the autonomic nervous system. In this study, Usui and colleagues administered CRF into the cerebral ventricles (i.c.v.) of anesthetized rats. Thirty and 60 min after CRF administration, the stellate, superior cervical and celiac ganglia were collected from euthanized animals and the ganglia were prepared for immunocytochemical localization of dopamine-β-hydroxylase (DβH; a marker for postganglionic sympathetic neurons) and c-Fos protein (a marker for activated neurons). The authors found that CRF treatment increased c-Fos immunoreactivity in the nuclei of neurons in the stellate and celiac ganglia but not in the superior cervical ganglion. The stimulatory effect was most prominent in the celiac
2
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
ganglion. These data show that central CRF does not produce a generalized autonomic activation but instead produces a selective activation of sympathetic preganglionic neurons that supply specific sympathetic ganglia and perhaps organ specific subsets of neurons within those activated ganglia. Commentary Stress produces a wide variety of responses that involve multiple organ systems. The autonomic nervous system is a major player in the stress response and there are many autonomic manifestations of stress. These include changes in cardiovascular, respiratory tract, gastrointestinal tract and urinary tract functions. CRF is released from hypothalamic neurons into the hypophyseal portal blood supply of the pituitary gland where it stimulates release of adrenocorticotropin which then stimulates release of cortisol from the adrenal cortex. However, increased activity of CRF neurons controlling central autonomic outflow is also part of the stress response. Usui et al. show that centrally administered CRF selectively activates subsets of sympathetic preganglionic neurons. This results in activation of neurons in the stellate ganglion (which supply the heart and lungs) and the celiac ganglion (which supply splanchnic organs) but not the superior cervical ganglion (which largely supplies targets in the head and neck region). These data indicate that central CRF does not produce an overall autonomic arousal but instead there is some selectivity in the pathways activated by CRF. It is also interesting that central CRF did not activate all neurons within the stellate or celiac ganglia. It is not likely that insufficient CRF was administered to cause maximum activation as the authors used 1.5 and 3.0 nM doses. The high dose did not produce a greater neuronal activation than the lower dose. Also, there was no difference in the number of neurons activated at 60 and 120 min post CRF administration, indicating that sufficient time was allowed for maximum activation of the CRF sensitive pathways. These data indicate that there are specific sympathetic postganglionic neurons targeted by CRF sensitive preganglionic pathways. It will be important to identify the specific subsets of neurons activated by CRF as this will provide new details about the organization of the autonomic nervous system and its specific role in the stress response.
Metzger, M., Bareiss, P.M., Danker, T., Wagner, S., Hennenlotter, J., Guenther, E., Obermayr, F., Stenzl, A., Koenigsrainer, A., Skutella, T., Just, L. (Tuebingen, Germany). 2009. Expansion and differentiation of neural progenitors derived from the human adult enteric nervous system. Gastroenterology. Article summary There are a number of gastrointestinal motility and secretory disorders that result from congenital defects in the development and maturation of the enteric nervous system. These defects include Hirschprung's disease and chronic intestinal idiopathic pseudo-obstruction. Surgery is the only treatment and this does not always result in satisfactory outcomes. There is a need for the development of other treatments and transplantation of neural stem cells holds some promise. The work by Metzger and colleagues shows that it may be possible to isolate neural stem and progenitor cells from the enteric nervous system of the adult gastrointestinal tract and these isolated cells could potentially be used in cell-based therapies for developmental abnormalities in the enteric nervous system. Human enteric spheroids were generated from explants isolated from surgical specimens of the small and large intestine that had been removed from adult patients (26– 84 years old) undergoing surgery for cancer, diverticulitis and bladder reconstruction. The authors found that the enterospheroids could be
expanded in culture and spheroid-derived cells could be differentiated into neuronal subtypes after incubation with conditioned medium obtained from murine embryonic gut cultures. These neuronal subtypes included nitrergic, cholinergic and serotonergic neurons. Glial cells could also be identified. Whole cell voltage-clamp studies revealed that differentiated neurons expressed functional voltage-gated sodium and potassium channels. In some experiments, enterospheres were grafted to hindgut segments obtained from E11 day mouse embryos Enteric neurons have not populated the hind gut at this time. Individual cells migrated from the grafted enterosphere into aganglionic explants of mouse intestine and they expressed neuronal markers and had neurone-like extensions. These data indicate that it may be possible to harvest multipotent progenitor cells from the human intestine and that these cells can differentiate into subtypes of enteric neurons or glia. It may be possible to use these enterospheres in cell-based therapies of gastrointestinal motility and secretory disorders that are due to defects in the maturation of the enteric nervous system. Commentary Chronic gastrointestinal motility disorders are very common and unfortunately there are few effective drug treatments for these problems. In most of the patients suffering from these problems, there is no obvious defect in the nerve supply of the gut. However, there are a smaller number of patients who suffer from motility, secretory and absorptive disorders that are based on clear deficits in the nerve supply of the gut. The best known of these is Hirschprung's disease in which the developing enteric nervous system fails to populate the distal colon. Chronic intestinal pseudo-obstruction (CIPO) is a rare but extremely debilitating motility disorder that appears to be due to deficits in the enteric nervous system supplying portions of the small intestine. There are currently no effective treatments of CIPO other than to resect the paralyzed portion of the intestine. The work of Metzger and colleagues may provide some hope for these patients. They have demonstrated the feasibility of isolating and growing neural/glial progenitor cells from adult intestine and that these can be induced to differentiate into subtypes of enteric neurons or glial cells under appropriate conditions. The great advantage of their approach is that specimens of adult human intestine are the source material of which there is a ready supply. Resection of human intestine is performed for a variety of reasons and these tissues are readily available at most academic medical centers. It remains to be seen if these progenitor cells can be induced to differentiate into the full range of neuronal phenotypes present within the intestinal wall and whether they perform the functions of these neurons in re-populated tissues. Can they form synapses with other neurons? Can they maintain appropriate polarity of projection or find appropriate targets (muscle, other neurons, blood vessels, epithelial cells). There are many remaining issues but this study provides important new information about a potential treatment strategy for developmental defects of the enteric nervous system.
Nogueiras, R., Pérez-Tilve, D., Veyrat-Durebex, C., Morgan, D.A., Varela, L., Haynes, W.G., Patterson, J.T., Disse, E., Pfluger, P.T., López, M., Woods, S.C., DiMarchi, R., Diéguez, C., Rahmouni, K., Rohner-Jeanrenaud, F., Tschöp, M.H. (Cincinnati, OH USA). 2009. Direct control of peripheral lipid deposition by CNS GLP-1 receptor signaling is mediated by the sympathetic nervous system and blunted in diet-induced obesity. J Neurosci 29, 5916–5925. Article summary Glucagon-like peptide 1 (GLP-1) is a gut hormone released in response to nutrient absorption. GLP-1 is released into the circulation and it can cross the blood brain barrier. It may act on central nerve circuits involved in energy balance. GLP-1 is also found in neurons in the
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
brain that are involved in energy balance and food intake. The work of Nogueiras et al. investigated the contribution of central GLP-1 receptors in brain pathways regulating fat metabolism by adipocytes independent of feeding behavior. These investigators used lean and obese mice and mice lacking β-adrenergic receptors. They measured sympathetic nerve activity and the expression of enzymes involved in lipid metabolism. In wild-type mice, intracerebroventricular (i.c.v.) infusion of a GLP-1 analog decreased lipid storage in white adipose tissue and this effect was independent from food intake. This effect of i.c.v. GLP-1 was reduced in obese mice and in mice with β1, β2 and β3 adrenergic receptors knocked out. β-receptors (in particular β1 and β2) are known to mediate the regulation of adipocyte lipid metabolism by the sympathetic nervous system. Recordings of the activity of sympathetic nerves supplying white adipose tissue (epididymal fat pad) showed that i.c.v. GLP-1 produced a concentration dependent increase in sympathetic nerve activity. Therefore, the CNS GLP-1 control of adipocyte lipid metabolism appears to be mediated, at least in part, by the sympathetic nervous system and is independent of parallel changes in food intake and body weight. Importantly, the CNS GLP-1 system loses the capacity to modulate adipocyte metabolism in obese states, suggesting an obesity-induced adipocyte resistance to CNS GLP-1. Commentary Obesity is a major public health issue that is likely to become more serious in the future. While poor lifestyle decisions are a major contributor to obesity, there are physiological contributions as well. Understanding the physiological processes that control appetite, fat deposition and energy utilization can only help to understand the causes of obesity and also help to develop preventative treatment strategies. It is well established that activation of the sympathetic nervous system can mobilize fat stores and increase circulating free fatty acids however not all of the mechanisms responsible for sympathetic activation under different physiological conditions are known. GLP-1 is a hormone and central neurotransmitter involved in regulation of energy homeostasis. GLP-1 also regulates appetite. This paper demonstrates that GLP-1 is linked to central sympathetic activation resulting in fat mobilization from white adipose tissue. This response is mediated by β1 and β2 adrenergic receptors expressed by adipocytes. Perhaps the most important finding is that GLP-1 induced activation of the sympathetic nervous system and fat mobilization is impaired in diet-induced obesity at least in mice. Restoration of this mechanism in obese individuals might be a useful approach to the treatment of diet-induced obesity.
Ajmo Jr., C.T., Collier, L., Leonardo, C.C., Hall, A.A., Green, S.M., Womble, T.A., Cuevas, J., Alison, A.E., Pennypacker, K.R. (Tampa, Fl, USA). 2009. Blockade of adrenoreceptors inhibits the splenic response to stroke. Exp Neurol 2009, 218: 47–55. Article summary Previously these authors have demonstrated, in the permanent middle cerebral artery occlusion (MCAO) model of stroke, that splenectomy 2 weeks prior to MCAO reduced the size of the infarction by N80%, highlighting the importance of this immune organ in the generation of secondary injury. They have also demonstrated that MCAO is followed by a reduction in spleen size and that this is associated with an increase in circulating regulatory T cells and macrophages. This study examines whether circulating or nerve-released catecholamines regulate the splenic response to stroke. Rats either underwent splenic denervation 2 weeks prior to MCAO or received α-adrenoceptor antagonists carvedilol (blocks α- and β-receptors), prazosin (α1-receptors) or propranolol (β-receptors). Denervation (confirmed by a marked reduction in TH labeled axons) prior to MCAO did not alter infarct volume or
3
spleen size. Propranolol treatment also had no effects on these outcomes. Prazosin or carvedilol treatment prevented the reduction in spleen size but only carvedilol significantly reduced infarct volume. The authors conclude that blood borne catecholamines released from the adrenal medulla regulate the splenic response to stroke through the activation of both α- and β-receptors. Commentary A prominent inflammatory response occurs following both ischemic and hemorrhagic stroke, thereby exacerbating the injury. There is also growing evidence that systemic inflammation influences stroke outcome and that therapies may need to also attenuate systemic inflammation to be effective. Indeed, the beneficial effects of stem cell therapy may be mediated, at least in part, by its systemic antiinflammatory effects. For example the authors have previously demonstrated that systemic treatment with human umbilical cord blood cells after MCAO prevents the reduction in spleen size, promotes expression of anti-inflammatory cytokines in the spleen and reduces neutrophil infiltration at the infarct site in the MCAO model. In this paper, the authors have investigated the involvement of the sympathetic nervous system in triggering the response of the spleen to MCAO. The findings demonstrate that carvedilol, possibly through a combined blockade of α- and β-adrenoceptors, reduces secondary injury at the infarct site. As blockade of α-adrenoceptors alone did not limit infarct volume but prevented the stroke-induced reduction in spleen size, these two events can be dissociated. Instead the authors suggest a role for circulating catecholamines in mediating the pro-inflammatory effects produced by MCAO in the spleen but do not identify their site(s) of action. This contrasts with the known anti-inflammatory action of activating β2adrenoceptors expressed on innate and adaptive immune cells; activation of these receptors by nerve-released and circulating catecholamines is believed to contribute to post-stroke immune suppression. The importance of this paper is that it demonstrates that stroke therapies should not focus solely on the central nervous system, but also target the peripheral immune cell signaling that plays a key role in ischemic injury progression. This will require greater understanding of role the sympathetic nervous system plays in regulating immune function.
Bove, G.M. (Portland, ME, USA). Focal nerve inflammation induces neuronal signs consistent with symptoms of early complex regional pain syndromes. Exp Neurol (Epub, ahead of print, 2009). Article summary During the early (acute) phase of complex regional pain syndromes (CRPS) there is both severe pain and autonomic dysfunction in the affected limb. While CRPS can be caused by overt nerve injury (type II), it can also result from relatively minor trauma (type I). Inflammation has been implicated in the etiology of CRPS. The author hypothesizes that inflammation of a nerve proximal to the affected region may result in changes that are consistent with clinical CRPS. To investigate this possibility he used a rat model of neuritis (induced using Freund's complete adjuvant) and recorded the activity of single cutaneous sensory and sympathetic axons at a point distal to the site of inflammation (in sural nerve following induction of neuritis in the sciatic nerve). In control rats, no sensory axons had ongoing activity and mean frequency of action potential discharge in sympathetic axons was 2.26 ± 1.33 Hz (mean ± SD). In contrast, in rats with inflamed nerves, 27% of slowly conducting sensory axons (b4 m/s: including Aδ- and C-neurons) had ongoing activity after 3–4 days, and 50% had such activity after 7–8 days. The sympathetic axons neurons had lower frequencies of ongoing discharge following induction of
4
J.J. Galligan, J.A. Brock / Autonomic Neuroscience: Basic and Clinical 150 (2009) 1–4
neuritis (1.96 ± 1.19 Hz after 3–4 days, 1.48 ± 1.23 Hz after 7–8 days). These findings demonstrate that focal nerve inflammation of a nerve trunk is sufficient to cause changes in the activity of both sensory and sympathetic axons projecting to skin. Commentary The clinical features of CRPS are pain, impaired motor function, edema and abnormalities due to altered sympathetic nerve activity (changes in sweating and blood flow). Furthermore, in some cases, ‘coupling’ between the sympathetic and sensory nervous systems plays a role in maintaining ongoing pain, although the precise locus for this interaction is not known. There is growing evidence that the initial insult results in changes to the central nervous system that modify sensory, motor and autonomic function. In patients with acute CRPS I, Baron and colleagues provided evidence that activity of sympathetic vasoconstrictor neurons is unilaterally reduced, causing swelling and redness of the affected limb (see the papers Discussion). These investigators have also presented evidence that sympathetic nerve activity is reduced during chronic CRPS I, although during this phase skin temperature and blood flow are reduced indicating
considerable cutaneous vasoconstriction. This paradox may be explained by ‘supersensitivity’ of the vasculature induced by the lower levels of sympathetic nerve activation (like decentralizationinduced supersensitivity). Furthermore, reduced levels of ongoing sympathetic nerve activity have been implicated in the acquisition of adrenergic sensitivity by nociceptors. In this paper the investigator has for the first time investigated the effects of experimental neuritis on sympathetic nerve activity. In previous papers, he has demonstrated that focal inflammation with Freund's complete adjuvant induces axonal mechano-sensitivity of intact deep somatic afferent A- and C-fibers and that this ‘sensitization’ is almost absent in cutaneous afferents. However, the present study indicates that focal neuritis induces ectopic activity in many of the C- and slowly conducting Aδ-afferent fibers supplying skin of the rat hindpaw (up to 50%; likely to be mostly nociceptive). As has been suggested in CRPS, the centrally determined level of ongoing activity in cutaneous sympathetic vasoconstrictor axons (baroreceptor sensitive) was concomitantly reduced. These changes would be expected to cause pain and contribute to hyperemia and swelling, perceived and expressed in the territory supplied by these axons (i.e. the hindpaw) and this now needs to be investigated.