Exercise participation barrier prevalence and association with exercise participation status in individuals with spinal cord injury.

STUDY DESIGN: Pass-code protected web survey. OBJECTIVES: Defining exercise participation barrier prevalence and association with exercise participation status in adults with spinal cord injury (SCI). SETTING: World-wide web. METHODS: Individuals ≥18 years with ShCI in the United States completed a pass-code protected website survey (N=180). Odds ratios (OR) and OR 95% confidence interval (95% CI) assessed association between barrier presence and exercise participation. RESULTS: No differences existed between exercisers and non-exercisers with respect to age, gender, injury level, injury duration, education level, or employment status. A larger percentage of non-exercisers reported household annual incomes <$7,500. The five most prevalent barriers were not associated with participation status (all OR 95% CI included 1). Low prevalence (≤13%) characterized four of the five barriers most strongly related to being a non-exerciser. Identifying too lazy, too difficult, or no interest as a barrier decreased odds of being an exerciser by 86%, 83%, and 71%, respectively. Not liking exercise decreased the odds of being an exerciser by 90%. CONCLUSION: Highly prevalent barriers were not associated with exercise participation status, whereas low prevalence barriers were strongly related to being a non-exerciser. Internal barriers had the strongest association with exercise participation status. The possible association between socioeconomic factors and exercise participation may be underappreciated. The most effective interventions to increase exercise participation may be multifocal approaches to enhance internal perceptions about and motivation to exercise, increase knowledge of how and where to exercise, while also reducing program and transportation financial costs.

United States (US) multi-center study to assess the validity and reliability of the Spinal Cord Independence Measure (SCIM III).

STUDY DESIGN: Multi-center, prospective, cohort study. OBJECTIVES: To assess the validity and reliability of the Spinal Cord Independence Measure (SCIM III) in measuring functional ability in persons with spinal cord injury (SCI). SETTING: Inpatient rehabilitation hospitals in the United States (US). METHODS: Functional ability was measured with the SCIM III during the first week of admittance into inpatient acute rehabilitation and within one week of discharge from the same rehabilitation program. Motor and sensory neurologic impairment was measured with the American Spinal Injury Association Impairment Scale. The Functional Independence Measure (FIM), the default functional measure currently used in most US hospitals, was used as a comparison standard for the SCIM III. Statistical analyses were used to test the validity and reliability of the SCIM III. RESULTS: Total agreement between raters was above 70% on most SCIM III tasks and all κ-coefficients were statistically significant (P<0.001). The coefficients of Pearson correlation between the paired raters were above 0.81 and intraclass correlation coefficients were above 0.81. Cronbach's-α was above 0.7, with the exception of the respiration task. The coefficient of Pearson correlation between the FIM and SCIM III was 0.8 (P<0.001). For the respiration and sphincter management subscale, the SCIM III was more responsive to change, than the FIM (P<0.0001). CONCLUSION: Overall, the SCIM III is a reliable and valid measure of functional change in SCI. However, improved scoring instructions and a few modifications to the scoring categories may reduce variability between raters and enhance clinical utility.

Site-directed neovessel formation in vivo.

Angiogenesis is an important component of organogenesis and wound repair and occurs during the pathology of oncogenesis, atherogenesis, and other disease processes. Thus, it is important to understand the physiological mechanisms that control neovascularization, especially with methods that permit the molecular dissection of the phenomenon in vivo. Heparin-binding growth factor-1 was shown to bind to collagen type I and type IV. When complexed with gelatin, heparin-binding growth factor-1 can induce neovascularization at polypeptide concentrations that are consistent with the biological activity of the mitogen in vitro. The adsorption strategy induces rapid blood vessel formation at and between organ- and tissue-specific sites and permits recovery of the site-specific implant for examination and manipulation by molecular methods.

Outcome measures in spinal cord injury: recent assessments and recommendations for future directions.

STUDY DESIGN: Review by the spinal cord outcomes partnership endeavor (SCOPE), which is a broad-based international consortium of scientists and clinical researchers representing academic institutions, industry, government agencies, not-for-profit organizations and foundations. OBJECTIVES: Assessment of current and evolving tools for evaluating human spinal cord injury (SCI) outcomes for both clinical diagnosis and clinical research studies. METHODS: a framework for the appraisal of evidence of metric properties was used to examine outcome tools or tests for accuracy, sensitivity, reliability and validity for human SCI. RESULTS: Imaging, neurological, functional, autonomic, sexual health, bladder/bowel, pain and psychosocial tools were evaluated. Several specific tools for human SCI studies have or are being developed to allow the more accurate determination for a clinically meaningful benefit (improvement in functional outcome or quality of life) being achieved as a result of a therapeutic intervention. CONCLUSION: Significant progress has been made, but further validation studies are required to identify the most appropriate tools for specific targets in a human SCI study or clinical trial.

Increasing thermal stress for tropical coral reefs: 1871-2017.

Tropical corals live close to their upper thermal limit making them vulnerable to unusually warm summer sea temperatures. The resulting thermal stress can lead to breakdown of the coral-algal symbiosis, essential for the functioning of reefs, and cause coral bleaching. Mass coral bleaching is a modern phenomenon associated with increases in reef temperatures due to recent global warming. Widespread bleaching has typically occurred during El Niño events. We examine the historical level of stress for 100 coral reef locations with robust bleaching histories. The level of thermal stress (based on a degree heating month index, DHMI) at these locations during the 2015-2016 El Niño was unprecedented over the period 1871-2017 and exceeded that of the strong 1997-1998 El Niño. The DHMI was also 5 times the level of thermal stress associated with the 'pre-industrial', 1877-1878, El Niño. Coral reefs have, therefore, already shown their vulnerability to the modest (~0.92 °C) global warming that has occurred to date. Estimates of future levels of thermal stress suggest that even the optimistic 1.5 °C Paris Agreement target is insufficient to prevent more frequent mass bleaching events for the world's reefs. Effectively, reefs of the future will not be the same as those of the past.

Gastroschisis in the United States 1988-2003: analysis and risk categorization of 4344 patients.

OBJECTIVE: Gastroschisis is a rare congenital abdominal wall defect through which intraabdominal organs herniate and it requires surgical management soon after birth. The objectives of this study were to profile patient characteristics of this anomaly utilizing data from two large national databases and to validate previous risk stratification categories of infants born with this condition. METHODS: An analysis was performed using 13 years of the National Inpatient Sample database (1988-1996, 1998, 1999, 2001, 2002) and 3 years of the Kids' Inpatient Database (1997, 2000, 2003). These combined databases contain information from nearly 93 million discharges in the United States. Infants with gastroschisis were identified by International Classification of Disease-9 procedure code 54.71 (repair of gastroschisis) and an age at admission of <8 days. Variables of gender, race, geographic region, co-existing diagnoses, length of stay, hospital charges adjusted to 2005 dollars, complications and inpatient mortality were collected from the databases. Infants were divided into simple and complex categories based on the absence or presence of intestinal atresia, stenosis, perforation, necrosis or volvulus. Comparisons between groups were performed using Pearson's chi (2) for categorical outcomes and the Kruskal-Wallis test for non-normally distributed continuous variables. RESULTS: A total of 4344 infants with gastroschisis were identified. These were comprised of 44.0% female infants (n=1910), 46.4% male infants (n=2017) whereas 9.6% were not reported (n=415). Racial analysis showed the largest subset being white in 40.9% of infants (n=1775) with Hispanic infants being the next highest group reported at 17.2% (n=745). Co-existing intestinal anomalies were the most common, affecting 9.9% (n=429) infants, whereas certain cardiac (6.8%, n=294) and pulmonary (1.7%, n=72) conditions were also identified. Simple gastroschisis represented 89.1% (n=3870) of the group whereas 10.9% (n=474) were complex in nature. Simple and complex patients differed in median length of stay (28 vs 67 days, P<0.01), inpatient mortality (2.9 vs 8.7%, P<0.01) and median inflation-adjusted hospital charges (90,788 dollars vs 197,871 dollars, P<0.01). CONCLUSIONS: These data represent a national analysis of the largest group of infants with gastroschisis to date which further aids the characterization and understanding of this serious congenital condition.

The RNA-binding protein HuD is required for GAP-43 mRNA stability, GAP-43 gene expression, and PKC-dependent neurite outgrowth in PC12 cells.

The RNA-binding protein HuD binds to a regulatory element in the 3' untranslated region (3' UTR) of the GAP-43 mRNA. To investigate the functional significance of this interaction, we generated PC12 cell lines in which HuD levels were controlled by transfection with either antisense (pDuH) or sense (pcHuD) constructs. pDuH-transfected cells contained reduced amounts of GAP-43 protein and mRNA, and these levels remained low even after nerve growth factor (NGF) stimulation, a treatment that is normally associated with protein kinase C (PKC)-dependent stabilization of the GAP-43 mRNA and neuronal differentiation. Analysis of GAP-43 mRNA stability demonstrated that the mRNA had a shorter half-life in these cells. In agreement with their deficient GAP-43 expression, pDuH cells failed to grow neurites in the presence of NGF or phorbol esters. These cells, however, exhibited normal neurite outgrowth when exposed to dibutyryl-cAMP, an agent that induces outgrowth independently from GAP-43. We observed opposite effects in pcHuD-transfected cells. The GAP-43 mRNA was stabilized in these cells, leading to an increase in the levels of the GAP-43 mRNA and protein. pcHuD cells were also found to grow short spontaneous neurites, a process that required the presence of GAP-43. In conclusion, our results suggest that HuD plays a critical role in PKC-mediated neurite outgrowth in PC12 cells and that this protein does so primarily by promoting the stabilization of the GAP-43 mRNA.

Initial UK experience with transversus abdominis muscle release for posterior components separation in abdominal wall reconstruction of large or complex ventral hernias: a combined approach by general and plastic surgeons.

Introduction Large, complicated ventral hernias are an increasingly common problem. The transversus abdominis muscle release (TAMR) is a recently described modification of posterior components separation for repair of such hernias. We describe our initial experience with TAMR and sublay mesh to facilitate abdominal wall reconstruction. Methods The study is a retrospective review of patients undergoing TAMR performed synchronously by gastrointestinal and plastic surgeons. Results Twelve consecutive patients had their ventral hernias repaired using the TAMR technique from June 2013 to June 2014. Median body mass index was 30.8kg/m2 (range 19.0-34.4kg/m2). Four had a previous ventral hernia repair. Three had previous laparostomies. Four had previous stomas and three had stomas created at the time of the abdominal wall reconstruction. Average transverse distance between the recti was 13cm (3-20cm). Median operative time was 383 minutes (150-550 minutes) and mesh size was 950cm2 (532-2400cm2). Primary midline fascial closure was possible in all cases, with no bridging. Median length of hospital stay was 7.5 days (4-17 days). Three developed minor abdominal wall wound complications. At median review of 24 months (18-37 months), there have been no significant wound problems, mesh infections or explants, and none has developed recurrence of their midline ventral hernia. Visual analogue scales revealed high patient satisfaction levels overall and with their final aesthetic appearance. Conclusions We believe that TAMR offers significant advantages over other forms of components separation in this patient group. The technique can be adopted successfully in UK practice and combined gastrointestinal and plastic surgeon operating yields good results.

Coral recovery in the central Maldives archipelago since the last major mass-bleaching, in 1998.

Increasing frequency and severity of disturbances is causing global degradation of coral reef ecosystems. This study examined temporal changes in live coral cover and coral composition in the central Maldives from 1997 to 2016, encompassing two bleaching events, a tsunami, and an outbreak of Acanthaster planci. We also examined the contemporary size structure for five dominant coral taxa (tabular Acropora, Acropora muricata, Acropora humilis, Pocillopora spp, and massive Porites). Total coral cover increased throughout the study period, with marked increases following the 1998 mass-bleaching. The relative abundance of key genera has changed through time, where Acropora and Pocillopora (which are highly susceptible to bleaching) were under-represented following 1998 mass-bleaching but increased until outbreaks of A. planci in 2015. The contemporary size-structure for all coral taxa was dominated by larger colonies with peaked distributions suggesting that recent disturbances had a disproportionate impact on smaller colonies, or that recruitment is currently limited. This may suggest that coral resilience has been compromised by recent disturbances, and further bleaching (expected in 2016) could lead to highly protracted recovery times. We showed that Maldivian reefs recovered following the 1998 mass-bleaching event, but it took up to a decade, and ongoing disturbances may be eroding reef resilience.

The molecular basis of lung morphogenesis.

To form a diffusible interface large enough to conduct respiratory gas exchange with the circulation, the lung endoderm undergoes extensive branching morphogenesis and alveolization, coupled with angiogenesis and vasculogenesis. It is becoming clear that many of the key factors determining the process of branching morphogenesis, particularly of the respiratory organs, are highly conserved through evolution. Synthesis of information from null mutations in Drosophila and mouse indicates that members of the sonic hedgehog/patched/smoothened/Gli/FGF/FGFR/sprouty pathway are functionally conserved and extremely important in determining respiratory organogenesis through mesenchymal-epithelial inductive signaling, which induces epithelial proliferation, chemotaxis and organ-specific gene expression. Transcriptional factors including Nkx2.1, HNF family forkhead homologues, GATA family zinc finger factors, pou and hox, helix-loop-helix (HLH) factors, Id factors, glucocorticoid and retinoic acid receptors mediate and integrate the developmental genetic instruction of lung morphogenesis and cell lineage determination. Signaling by the IGF, EGF and TGF-beta/BMP pathways, extracellular matrix components and integrin signaling pathways also directs lung morphogenesis as well as proximo-distal lung epithelial cell lineage differentiation. Soluble factors secreted by lung mesenchyme comprise a 'compleat' inducer of lung morphogenesis. In general, peptide growth factors signaling through cognate receptors with tyrosine kinase intracellular signaling domains such as FGFR, EGFR, IGFR, PDGFR and c-met stimulate lung morphogenesis. On the other hand, cognate receptors with serine/threonine kinase intracellular signaling domains, such as the TGF-beta receptor family are inhibitory, although BMP4 and BMPR also play key inductive roles. Pulmonary neuroendocrine cells differentiate earliest in gestation from among multipotential lung epithelial cells. MASH1 null mutant mice do not develop PNE cells. Proximal and distal airway epithelial phenotypes differentiate under distinct transcriptional control mechanisms. It is becoming clear that angiogenesis and vasculogenesis of the pulmonary circulation and capillary network are closely linked with and may be necessary for lung epithelial morphogenesis. Like epithelial morphogenesis, pulmonary vascularization is subject to a fine balance between positive and negative factors. Angiogenic and vasculogenic factors include VEGF, which signals through cognate receptors flk and flt, while novel anti-angiogenic factors include EMAP II.

Striatonigral projection neurons: a retrograde labeling study of the percentages that contain substance P or enkephalin in pigeons.

Two largely separate populations of neuropeptide-containing striatonigral projection neurons have been distinguished in pigeons, one population whose neurons contain substance P (SP) and dynorphin (DYN) and a second population whose neurons contain enkephalin (ENK) (Reiner, '86a; Anderson and Reiner, '90a). In the present study, we investigated the abundance of these two types of neurons relative to all striatonigral projection neurons by combining retrograde labeling by the fluorescent dye fluorogold with immunofluorescence labeling for SP and ENK. Pigeons received large intranigral injections of fluorogold to retrogradely label the striatonigral projection neurons, and several days later they were treated with colchicine (32 hours before transcardial perfusion). Adjacent series of sections through the basal ganglia were labeled for SP and ENK using immunofluorescence techniques. The tissue was examined using fluorescence microscopy and the percentages of retrogradely labeled neurons containing either SP or ENK were quantified. We found that 85-95% of the fluorogold-labeled striatonigral neurons were SP+, whereas only 1-4% were ENK+. Thus the majority of striatonigral projection neurons in pigeons appear to contain SP, whereas a small percentage contain ENK. Only a small percentage of striatonigral neurons did not contain either. Since striatal projection neurons also contain GABA (Reiner, '86b), the present results suggest that a high percentage of striatonigral projection neurons coexpress SP, DYN and GABA, whereas a small fraction coexpress ENK and GABA. The available data are consistent with the conclusion that this is true in reptilian and mammalian species as well.

Distribution and relative abundance of neurons in the pigeon forebrain containing somatostatin, neuropeptide Y, or both.

Immunohistochemical studies in several mammalian species and in red-eared turtles have shown that somatostatin (SS) and neuropeptide Y (NPY) co-occur in a substantial proportion of the telencephalic neurons containing either. To explore further the possibility that telencephalic neurons co-containing SS and NPY may be evolutionarily conserved among amniotes, we determined the distribution and co-occurrence of SS and NPY in forebrain neurons in pigeons. Single-label immunohistochemical studies revealed the presence of overlapping populations of SS+ neurons and NPY+ neurons in most of the major subdivisions of the telencephalon. Double-label immunofluorescence studies revealed that in subdivisions of the telencephalon that are comparable to mammalian cortex (i.e., those dorsal and lateral to the basal ganglia), the vast majority of NPY+ neurons were also SS+, whereas a major and regionally variable percentage of the SS+ neurons were not NPY+. In contrast, within the basal telencephalon (including the basal ganglia and several other structures) neurons labeled only for NPY or only SS were more abundant than those containing both neuropeptides. Outside the telencephalon, the only forebrain cell group containing neurons in which SS and NPY were co-localized was in the lateral hypothalamus. A series of double- and triple-label immunohistochemical studies was undertaken to determine the extent of co-occurrence of SS and NPY in striatal neurons and the relationship of these neurons to striatal neurons containing other neuropeptides. In addition, immunohistochemical single- and double-label techniques were employed in conjunction with retrograde-labeling by fluorogold to determine the projections of SS+ and NPY+ striatal neurons. The results indicate that: 1) a population of striatal interneurons containing both SS and NPY exists in pigeons and constitutes approximately the same fraction of all striatal neurons as reported in mammals, 2) neurons containing NPY (but not SS) form a second, larger population of striatal interneurons, 3) neurons containing SS (but not NPY) form a third population of striatal interneurons that is approximately half as abundant as the NPY+ interneuron population, and 4) one-third of the substance P-containing striatonigral projection neurons also contain SS. The existence in pigeons of a major population of neurons containing both SS and NPY throughout the telencephalon, the existence of a population of neurons containing only SS in cortex-equivalent parts of the telencephalon, and the existence of a population of interneurons containing only NPY in the striatum is consistent with findings in mammals and turtles.(ABSTRACT TRUNCATED AT 400 WORDS)

Extensive co-occurrence of substance P and dynorphin in striatal projection neurons: an evolutionarily conserved feature of basal ganglia organization.

A number of different neuroactive substances have been found in striatal projection neurons and in fibers and terminals in their target areas, including substance P (SP), enkephalin (ENK), and dynorphin (DYN). In a preliminary report on birds and reptiles, we have suggested that SP and DYN are to a large extent found in the same striatal projection neurons and that ENK is found in a separate population of striatal projection neurons. In the present study, we have examined this issue in more detail in pigeons and turtles. Further, we have also explored this issue in rats to determine whether this is a phylogenetically conserved feature of basal ganglia organization. Simultaneous immunofluorescence double-labeling procedures were employed to explore the colocalization of SP and DYN, SP and ENK, and ENK and DYN in striatal neurons and in striatal, nigral, and pallidal fibers in pigeons, turtles, and rats. To guard against possible cross-reactivity of DYN and ENK antisera with each others' antigens, separate double-label studies were carried out with several different antisera that were specific for DYN peptides (e.g., dynorphin A 1-17, dynorphin B, leumorphin) or ENK peptides (leucine-enkephalin, metenkephalin-arg6-gly7-leu8, methionine-enkephalin-arg6-phe7). The results showed that SP and DYN co-occur extensively in specific populations of striatal projection neurons, whereas ENK typically is present in different populations of striatal projection neurons. In pigeons, 95-99% of all striatal neurons containing DYN were found to contain SP and vice versa. In contrast, only 1-3% of the SP+ striatal neurons and no DYN neurons contained ENK. Similarly, in turtles, greater than 75% of the SP+ neurons were DYN+ and vice versa, whereas ENK was observed in fewer than 5% of the SP+ neurons and 2% of the DYN+ neurons. Finally, in rats, more than 70% of the SP+ neurons contained DYN and vice versa, but ENK was found in only 5% of the SP+ neurons and in none of the DYN+ perikarya. Fiber double-labeling in the striatum and its target areas (the pallidum and substantia nigra) was also consonant with these observations in pigeons, turtles, and rats. These results, in conjunction with studies in cats by M.-J. Besson, A.M. Graybiel, and B. Quinn (1986; Soc Neurosci. Abs. 12:876) strongly indicate that the co-occurrence of SP and DYN in large numbers of striatonigral and striatopallidal projection neurons in a phylogenetically widespread, and therefore evolutionarily conserved, feature of basal ganglia organization.(ABSTRACT TRUNCATED AT 400 WORDS)

Ultrastructural single- and double-label immunohistochemical studies of substance P-containing terminals and dopaminergic neurons in the substantia nigra in pigeons.

The vast majority of striatonigral projection neurons in pigeons contain substance P (SP), and the vast majority of SP-containing fibers terminating in the substantia nigra arise from neurons in the striatum. To help clarify the role of striatonigral projection neurons, we conducted electron microscopic single- and double-label immunohistochemical studies of SP+ terminals and/or dopaminergic neurons (labeled with either anti-dopamine, DA, or anti-tyrosine hydroxylase, TH) in pigeons to determine: (1) the synaptic organization of SP+ terminals, (2) the synaptic organization of TH+ perikarya and/or dendrites, and (3) the synaptic relationship between SP+ terminals and TH+ neurons in the substantia nigra. Tissue single-labeled for SP revealed numerous SP+ terminals contacting thin unlabeled dendrites in the substantia nigra, but few SP+ terminals were observed contacting perikarya or large-diameter dendrites. SP+ terminals contained round, densely packed, clear vesicles, and often contained one or more dense-core vesicles. Synaptic junctions between SP+ terminals and their targets were more often symmetric (86%) than asymmetric. In tissue single-labeled for DA, we observed few terminals contacting DA+ perikarya, whereas terminals contacting DA+ dendrites were more abundant. Terminals contacting DA+ structures comprised at least four different morphologically distinct types based on the morphology of the clear synaptic vesicles and the type of synaptic junction. One type of terminal contained round clear vesicles and made symmetric synapses, and thus resembled the predominant type of SP+ terminal. The second type contained round clear vesicles and made asymmetric synapses, the third type contained medium-size pleomorphic clear vesicles and made symmetric synapses, and the fourth type contained small pleomorphic clear vesicles and made symmetric synapses. The presence of contacts between SP+ terminals and dopaminergic dendrites in the substantia nigra was directly demonstrated in tissue double-labeled for SP (by the peroxidase-antiperoxidase procedure, or PAP, with diaminobenzidine) and TH (by either the silver-intensified immunogold procedure or the PAP procedure with benzidine dihydrochloride). SP+ terminals commonly contacted thin TH+ dendrites in the substantia nigra, but few SP+ terminals contacted large-diameter TH+ dendrites or perikarya. Synapses between SP+ terminals and TH+ neurons were always symmetric. TH+ dendrites also were contacted by terminals not labeled for SP, which were more abundant than were SP+ terminals. Non-TH+ neurons were also contacted by both SP+ terminals and non-SP+ terminals.(ABSTRACT TRUNCATED AT 400 WORDS)

An ultrastructural double-label immunohistochemical study of the enkephalinergic input to dopaminergic neurons of the substantia nigra in pigeons.

Electron microscopic immunohistochemical double-label studies were carried out in pigeons to characterize the ultrastructural organization and postsynaptic targets of enkephalinergic (ENK+) striatonigral projection. ENK+ terminals in the substantia nigra were labeled with antileucine-enkephalin antiserum by using peroxidase-antiperoxidase methods, and dopaminergic neurons were labeled with anti-tyrosine hydroxylase antiserum by using silver-intensified immunogold methods. ENK+ terminals on dopaminergic neurons were equal in abundance to ENK+ terminals on nondopaminergic neurons, although the former were typically somewhat smaller than the latter (mean size: 0.50 vs. 0.75 micron, respectively). ENK+ terminals were evenly distributed on the cell bodies and dendrites of dopaminergic neurons, and they were evenly distributed on dendrites but rare on perikarya of nondopaminergic neurons. Transection of the basal telencephalic output revealed that 75% of the nigral ENK+ terminals were of basal telencephalic origin. These telencephalic ENK+ terminals included over 80% of those smaller than 0.80 micron on dopaminergic neurons and smaller than 1.0 micron on nondopaminergic neurons, and none greater than this in size. Both telencephalic and the nontelencephalic ENK+ nigral terminals made predominantly symmetric synapses on nigral neurons. Although the basal telencephalic ENK+ terminals uniformly targeted dendrites and perikarya, nontelencephalic ENK+ terminals seemed to avoid perikarya. The results indicate that ENK+ striatonigral neurons in birds may directly influence both dopaminergic and nondopaminergic neurons of the substantia nigra. Based on similar data for substance P-containing striatonigral terminals, the roles of enkephalin and substance P in influencing nigral dopaminergic neurons may differ slightly, as they appear to target preferentially different portions of dopaminergic neurons. The overall results in pigeons are similar to those for ENK+ terminals in the ventral tegmental area in rats, suggesting that the synaptic organization of the ENK+ input to the tegmental dopaminergic cell fields is similar in mammals and birds.

Light and electron microscopic immunohistochemical study of dopaminergic terminals in the striatal portion of the pigeon basal ganglia using antisera against tyrosine hydroxylase and dopamine.

A dopaminergic projection from the midbrain to the striatal portion of the basal ganglia is present in reptiles, birds, and mammals. Although the ultrastructure of these fibers and terminals within the striatum has been studied extensively in mammals, little information is available on the ultrastructure of this projection in nonmammals. In the present study, we used immunohistochemical labeling with antibodies against tyrosine hydroxylase (TH) or dopamine (DA) to study the dopaminergic input to the striatal portion of the basal ganglia in pigeons (i.e., lobus parolfactorius and paleostriatum augmentatum). At the light microscopic level, the anti-TH and anti-DA revealed a similar abundance and distribution of numerous labeled fine fibers and varicosities within the striatum. In contrast, the use of an antidopamine beta-hydroxylase antiserum (which labels only adrenergic and noradrenergic terminals) labeled very few striatal fibers, which were restricted to visceral striatum. These results demonstrate that anti-TH mainly labels dopaminergic terminals in the striatum. At the electron microscopic level, the anti-TH and anti-DA antisera labeled numerous axon terminals within the striatum (15-20% of all striatal terminals). These terminals tended to be small (with an average length of 0.6 microns) and flattened, and their vesicles tended to be small (35-60 nm in diameter) and pleomorphic. About 50% of the terminals were observed to make synaptic contacts in the planes of section examined, and nearly all of these synaptic contacts were symmetric. Both TH+ and DA+ terminals typically contacted dendritic shafts or the necks of dendritic spines, but a few contacted perikarya. No clear differences were observed between TH+ and DA+ terminals within medial striatum (whose neurons project to the nigra in birds) or between TH+ and DA+ terminals within lateral striatum (whose neurons project to the pallidum in birds). In addition, no differences were observed between medial and lateral striata in either TH+ or DA+ terminals. Thus, there is no evident difference in pigeons between striatonigral and striatopallidal neurons in their dopaminergic innervation. Our results also indicate that the abundance, ultrastructural characteristics, and postsynaptic targets of the midbrain dopaminergic input to the pigeon striatum are highly similar to those in mammals. This anatomical similarity is consistent with the pharmacologically demonstrable similarity in the role of the dopaminergic input to the striatum in birds and mammals.

Differential distribution of exogenous BDNF, NGF, and NT-3 in the brain corresponds to the relative abundance and distribution of high-affinity and low-affinity neurotrophin receptors.

To evaluate effective means for delivering exogenous neurotrophins to neuron populations in the brain, we compared the distribution and transport of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and neurotrophin-3 (NT-3) following intracerebral delivery. Rats received an injection of radioiodinated or unlabeled neurotrophin into the lateral ventricle and were killed humanely after 1.5-24 hours. Other rats received continuous infusion of unlabeled neurotrophin into the lateral ventricle, the striatum, or the hippocampus for 3-14 days. The neurotrophins were detected by autoradiography or immunohistochemistry. There were striking differences between BDNF, NGF, and NT-3 in their penetration through brain tissue. These differences occurred regardless of the site or method of delivery, but were most pronounced following a bolus intracerebroventricular (ICV) injection. After ICV injection, NGF was widely distributed in tissues around the ventricles and at the surface of the brain, whereas the penetration of BDNF into brain tissue was distinctly less than that of NGF, and the penetration of NT-3 was intermediate. An ICV injection of NGF produced prominent but transient labeling of cells that contain the low-affinity NGF receptor, whereas an injection of BDNF prominently labeled the ventricular ependyma. During ICV infusion (12 micrograms/day), the distribution of BDNF was no greater than that observed after a 12-micrograms bolus injection. A sixfold increase in the amount of BDNF infused (72 micrograms/day) produced a more widespread distribution in the brain and doubled the depth of penetration into periventricular tissues near the cannula. Corresponding to their differences in penetration, NGF was retrogradely transported by basal forebrain cholinergic neurons after ICV or intrastriatal delivery, whereas NT-3 was transported by a few basal forebrain neurons after ICV delivery, and BDNF was rarely detected in neurons after ICV delivery. Delivery of BDNF directly to the striatum or the hippocampus labeled numerous neurons in nuclei afferent to these structures. In situ hybridization studies confirmed that the high-affinity BDNF receptor (TrkB) was much more widely expressed in neurons than was the high-affinity NGF receptor (TrkA). Moreover, mRNA for truncated forms of TrkB was expressed at high levels in the ependyma, the choroid epithelium, and the gray matter. It is likely that binding of BDNF to TrkB, which appears to be more abundant and ubiquitous than TrkA, restricts the diffusion of BDNF relative to that of NGF.

A pre-embedding triple-label electron microscopic immunohistochemical method as applied to the study of multiple inputs to defined tegmental neurons.

For many neural regions it is of interest to know the identity of the target structures of two different types of inputs to that neural region. Such studies require use of a triple-label immunohistochemical method to differentially label the class of target structure and the two types of input so that they can be visualized at the electron microscopic (EM) level. We describe here a procedure for combining three different markers (diaminobenzidine, benzidine dihydrochloride, and silver-intensified immunogold) for triple-label EM immunohistochemical pre-embedding labeling. All three markers are distinct at the LM and EM levels. An example of this approach as applied to studying striatal input to the ventral tegmental area is presented and the advantages of this approach are discussed.

Activation of the hypothalamic arcuate nucleus predicts the anorectic actions of ciliary neurotrophic factor and leptin in intact and gold thioglucose-lesioned mice.

Similar to leptin, ciliary neurotrophic factor (CNTF) suppresses appetite and selectively reduces body fat in leptin-deficient ob/ob mice. To assess the relative importance of specific regions of the hypothalamus in mediating these effects, we administered a CNTF analogue (CNTFAx15) or leptin to mice made obese by administration of gold thioglucose (GTG), which destroys a well-defined portion of the medial basal hypothalamus. CNTFAx15 treatment reduced appetite and body weight in obese GTG-lesioned C57BL/6 mice, whereas leptin failed to effect similar changes regardless of whether treatment was initiated before or after the lesioned mice had become obese. Because leptin does not reduce food intake or body weight in most forms of obesity (a condition termed 'leptin resistance'), we also investigated the actions of leptin in GTG-lesioned leptin-deficient (ob/ob) mice. By contrast to C57BL/6 mice, leptin treatment reduced food intake and body weight in GTG-lesioned ob/ob mice, although the effect was attenuated. To further compare the neural substrates mediating the anorectic actions of leptin and CNTF, we determined the patterns of neurone activation induced by these proteins in the hypothalamus of intact and GTG-lesioned mice by staining for phosphorylated signal transducer and activator of transcription 3 (pSTAT3). CNTFAx15 stimulated robust pSTAT3 signalling in neurones of the medial arcuate nucleus in both intact and lesioned C57BL/6 and ob/ob mice. Leptin administration stimulated pSTAT3 signalling in only a few neurones of the medial arcuate nucleus in intact or lesioned C57BL/6 mice, but elicited a robust response in intact or lesioned ob/ob mice. By contrast to CNTFAx15, leptin treatment also resulted in prominent activation of STAT3 in several areas of the hypothalamus outside the medial arcuate nucleus. This leptin-induced pSTAT3 signal was at least as prominent in intact and GTG-lesioned C57BL/6 mice as it was in ob/ob mice, and thus was not correlated with appetite suppression or weight loss. These results indicate that the medial arcuate nucleus is a key mediator of appetite suppression and weight loss produced by CNTF and leptin, whereas GTG-vulnerable regions play a role only in leptin-induced weight loss. Other regions of hypothalamus in which pSTAT3 signal is induced by leptin may regulate energy metabolism through mechanisms other than appetite reduction.

Thalamostriatal projection neurons in birds utilize LANT6 and neurotensin: a light and electron microscopic double-labeling study.

Based on its location, connectivity and neurotransmitter content, the dorsal thalamic zone in birds appears to be homologous to the intralaminar, midline, and mediodorsal nuclear complex in the thalamus of mammals. We investigated the neuroactive substances used by thalamostriatal projection neurons of the dorsal thalamic zone in the pigeon. Single-labeling experiments showed that many neurons in the dorsal thalamic zone are immunoreactive for neurotensin and the neurotensin-related hexapeptide, (Lys8,Asn9)NT(8-13) (LANT6). Double-labeling experiments, using the retrograde fluorescent tracer, FluoroGold, combined with fluorescence immunocytochemistry for either LANT6 or neurotensin, showed that neurotensin- and LANT6-containing neurons in the dorsal thalamic zone project to the striatum of the basal ganglia. Immunofluorescence double-labeling experiments showed that neurotensin and LANT6 are often (possibly always) co-expressed in neurons in the dorsal thalamic zone. Electron microscopic immunohistochemical double-labeling showed that LANT6 terminals in the striatum make asymmetric contacts with heads of spines labeled for substance P and heads of spines not labeled for substance P, suggesting that these terminals synapse with both substance P-containing and non-substance P-containing medium spiny striatal projection neurons. These findings indicate that LANT6 and neurotensin may be utilized as neurotransmitters in thalamostriatal projections in birds and raise the possibility that this may also be the case in other amniotes.