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The nucleus of tractus solitarius (NTS) is the main integration site for autonomic control. It receives afferent information from arterial baroreceptors, chemoreceptors, cardiac receptors, gastrointestinal receptors and respiratory receptors (1). Details on the neurochemical properties of NTS neurones have been established by many researchers. However information on the co-localisation of choline acetyltransferase (ChAT) and glutamic acid decarboxylase (GAD) immunoreactivity within the NTS has not been reported. This study examined the distribution of ChAT immunoreactivity and co-localisation with green fluorescent protein (GFP) in the NTS of transgenic mice (n=4) expressing GFP under control of the GAD67 promoter (GAD67-GFP mice) (2) using fluorescence immunohistochemistry (3). Four GAD67-GFP adult mice (mixed sexes) were injected intraperitoneally with 1 % Fluorogold. After 24-48 hours the mice were anaesthetised with 60 mg/kg sodium pentobarbitone and when the withdrawal reflex was absent, they were perfused transcardially with 4 % paraformaldehyde. The brainstems were postfixed with 4 % paraformaldehyde for 24 hours. Brainstems were sectioned (at 50 µm and 30 µm for double and triple labelling respectively). Serial sections were processed utilising fluorescent immunohistochemistry (IHC). One wild type adult mouse was also prepared in the same way as the GAD67-GFP mice. Sections were processed using diaminobenzidine IHC for ChAT to provide a permanent reaction product. ChAT immunoreactive (ChAT-IR) and GAD67-GFP immunoreactive (GAD67-GFP-IR) neurones were found in all NTS subnuclei. Moreover, co-localisation was observed in some NTS subnuclei; predominantly in the intermediate subnucleus (colocalised neurones per section were 6.83 ± 1.89 (mean ± SEM) for the right and 6.67 ± 2.19 for the left NTS column, n = 6 sections) and the central subnucleus (1.00 ± 0.68 (right NTS) and 2.17 ± 1.42 (left NTS)). Nitric oxide synthase immunoreactive (nNOS-IR) neurones in the central subnucleus are reported to be involved in oesophageal regulation (4). In our study, although nNOS-IR neurones were prominent in the central subnucleus, they did not exhibit co-localisation with either ChAT or GAD67-GFP. Although the absence of co-labelling in the central subnucleus with nNOS does not support a role for ChAT/GAD-IR neurones in oesophageal control, this possible function cannot be excluded at this time. Examination of other brainstem areas revealed co-localisation of ChAT and GAD67GFP immunoreactivity, including the area postrema, the reticular formation and the lateral paragigantocellular nucleus. Since there is limited information of co-localisation for ChAT and GAD immunoreactivity in the brainstem, it is worth studying further to investigate its roles in these neurones. [ABSTRACT FROM AUTHOR]

An abstract of the article "Co-localisation of the enzymes synthesising GABA and acetylcholine in the mouse spinal cord" by J. Gotts, I. J. Edwards, S. A. Deuchars and J. Deuchars is presented.

The central autonomic area (CAA) contains interneurones which directly influence sympathetic preganglionic neurones (Deuchars et al., 2005). Sensory afferent terminals are located around lamina X and form close appositions with interneurones (Hofstetter et al., 2005). CAA neurones have been identified as being premotor to somatic motor neurones (Stepien et al., 2010). This suggests that these interneurones may coordinate autonomic and motor outflow from the spinal cord and therefore, may receive inputs from primary afferents. This study aims to address how these interneurones fit into circuits involved in autonomic control. To investigate whether interneurones in CAA received direct inputs from myelinated primary afferents, immunohistochemistry for vesicular glutamate transporter 1 (VGLUT1) was conducted in GAD67-GFP mice anaesthetised with pentobarbital (60mg/kg; i.p) and transcardially perfused with 4% paraformaldehyde. Close appositions were observed between VGLUT1 primary afferent terminals and interneurones immunoreactive for GFP (GABAergic) or Choline Acetyltransferase. To determine how interneurones responded to stimulation at the dorsal root entry zone, 7-12 day old mice and 12-21 day old rats, of either sex, were anaesthetised by urethane (2g/kg; i.p). Transcardiac perfusion was followed by decapitation and 300µm transverse slices of lumbar and thoracic spinal cord were cut. Viable interneurones in laminae VII and X were identified and whole cell patch clamp recordings performed. A stimulating electrode was placed at the dorsal root entry zone to deliver stimuli (5-100 Hz) of 1 ms duration. The evoked excitatory postsynaptic potential (EPSP) amplitude was recorded and the latency to onset of the EPSP measured. The standard deviation (SD) of latency gave an indication of variability in latency (jitter). Values are means ± S.E.M, compared by 2 sample t-test. Mouse thoracic interneurones had an action potential threshold of -35.06 ± 1.99 mV, action potential amplitude of 49.52 ± 2.64 mV and afterhyperpolarisation amplitude of -19.45 ± 1.08 mv (n= 15). EPSP jitter was 2.31± 0.01 µs (n= 7). EPSPs followed high frequency tetanic stimulation, without failures indicating that they are likely elicited by monosynaptic primary afferents, and were significantly antagonised by 20 µM NBQX (an excitatory amino acid receptor antagonist) (7.68 ± 0.64 mV in control (n= 7) to 3.28 ± 0.45 mV (n= 7) in NBQX; p< 0.001). In rats, EPSP jitter was 4.23 ± 0.51 µs (n= 16) and EPSP amplitude was significantly reduced with NBQX (6.43 ± 0.04 mV (n= 16) in control to 0.42 ± 0.24 mV (n= 9) in 20 µM NBQX; p< 0.001). Post hoc analysis revealed extensive axon ramifications in both the dorsal and ventral horn. The direct action of primary afferents onto interneurones in the CAA may provide a avenue for the co-ordination of autonomic and motor activity. [ABSTRACT FROM AUTHOR]