Elsevier

Neuroscience

Volume 104, Issue 2, 10 May 2001, Pages 523-538
Neuroscience

Immunohistochemical profiles of spinal lamina I neurones retrogradely labelled from the nucleus tractus solitarii in rat suggest excitatory projections

https://doi.org/10.1016/S0306-4522(01)00071-9Get rights and content

Abstract

Three morphologically distinct types of lamina I neurones, fusiform, flattened and pyramidal, project from the spinal cord to the caudal part of the nucleus tractus solitarii in the rat, and may represent a pathway whereby peripheral stimuli can modify autonomic functions. The neurochemistry of these three types of projection neurones was investigated using retrograde neuronal tracing with cholera toxin B-subunit combined with dual and triple immunofluorescence labelling for different neuroactive substances. None of the lamina I neurones with immunoreactivity for GABA or glycine were found to project to the nucleus tractus solitarii, whereas high levels of glutamate immunoreactivity, which may indicate a glutamatergic phenotype, were found in 18.4% of fusiform, 9.6% of pyramidal and 2.1% of flattened projection neurones. Immunoreactivity for calbindin-D28K was present in 34.9% of fusiform cells, 18.3% of pyramidal cells and 10.5% of flattened cells, and nitric oxide synthase immunoreactivity was detected in 13.8% of fusiform cells, 1.1% of pyramidal cells and 4.2% of flattened cells that had projections to the nucleus tractus solitarii. Calbindin immunoreactivity was co-localised in major subpopulations of projection neurones of each morphological type that contained glutamate immunoreactivity, whereas co-localisation of nitric oxide synthase immunoreactivity in these neurones was relatively uncommon. The pyramidal cell was the only retrogradely labelled cell type found to be immunoreactive for substance P, but few (<5%) of these neurones were immunolabelled.

These data are consistent with the hypothesis that lamina I neurones projecting to the dorsal vagal complex are not inhibitory, and that some of them, belonging mostly to the fusiform and pyramidal types, may exert excitatory, glutamate- or substance P-mediated effects upon inhibitory interneurones in the nucleus tractus solitarii. Such excitatory pathways could be involved in the attenuation of the reflex control of blood pressure by both painful and innocuous peripheral stimuli, such as those arising in injury and exercise.

Section snippets

Surgical procedures

All procedures involving animal handling and usage were performed in accordance with the UK Animals (Scientific Procedures) Act 1986, European Council Directive 86/609/EEC, and the US Society for Neuroscience and NIH guidelines. Every effort was made to minimise any discomfort and suffering, and the minimum possible number of rats (n=11) was used to obtain reproducible and statistically significant results. Male Wistar rats (280–310 g; purpose bred at the University of Leeds) were anaesthetised

Injection sites

In eight animals used for the analysis, examination of sections through the medulla showed that the dense central cores of CTb immunoreactivity corresponding to the two injection sites lay within the commissural and medial subnuclei of the left NTS. The diffuse halos of immunoreactivity surrounding the points of injection filled the entire NTS, except for the most lateral parts at levels of the area postrema and more caudally, and extended to the ipsilateral dorsal motor nucleus of the vagus

Discussion

This study set out to define some of the putative neurotransmitters and other neuronal marker substances present in the different morphological categories of lamina I neurones that project to the NTS. GABA, glycine and glutamate were investigated, since these amino acid transmitters have previously been localised to neurones with a fusiform appearance,47., 63., 75., 81. and previous work has shown that this type accounts for the greatest number of marginal neurones projecting to the NTS in the

Acknowledgements

This work was supported by a British Heart Foundation project grant, a Medical Research Council JREI Award and a British Council “Treaty of Windsor” Travel Award. We are grateful to Dr Michael Conlon (Creighton University, Omaha, NE, USA), Dr Piers Emson (Babraham Institute, University of Cambridge, UK) and Dr David Pow (VTHRC, University of Queensland, Brisbane, Australia) for generously providing antisera used in this study.

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