Anthony D. Shafton

Neurons in the hypothalamic paraventricular nucleus send collaterals to the spinal cord and to the rostral ventrolateral medulla in the rat

The hypothalamic paraventricular nucleus (PVN) projects to the rostral ventrolateral medulla (RVLM) and to the intermediolateral cell column (IML) of the spinal cord. The present study determined whether the same neurons can innervate both regions. In each rat, two retrogradely-transported tracers, microspheres tagged with fluorescein or rhodamine, were injected into the left lower thoracic/upper lumbar IML (fluorescein) and into the pressor region of the left RVLM (rhodamine). In the PVN over 90% of the neurons labelled with either tracer were found ipsilateral to the injection site. Double labelled cells averaged almost one-third of the spinally-projecting cells in four of the five animals. In the remaining animal, there were few double-labelled cells. The results suggest that a population of PVN neurons innervates both the lower-thoracic/upper lumbar IML and the RVLM.

Evidence that stimulation of ghrelin receptors in the spinal cord initiates propulsive activity in the colon of the rat

Previous studies have failed to reveal an effect of the gastrointestinal peptide hormone ghrelin on colonic motility. In the present work, ghrelin was applied into the lumbo‐sacral spinal cord in the region of defecation control centres, and a synthetic ghrelin receptor agonist, CP464709, which crosses the blood–brain barrier, was applied intravenously or into the lumbo‐sacral cord. Both ghrelin and CP464709 elicited propulsive contractions and emptying of the colon in anaesthetized rats. In conscious rats, subcutaneous CP464709 caused fecal expulsion. The sites of action and nerve pathways involved in the stimulation of the colon by ghrelin receptor activation were investigated in anaesthetized rats. Intrathecal application of CP464709 at L6–S1, but not application at ponto‐medullary levels or to the thoracic spinal cord, elicited propulsive contractions. The stimulation evoked by intravenous CP464709 was prevented if the pelvic nerve outflows were severed, but not if the spinal cord was cut rostral to the defecation centre at L6–S3. The response was also blocked by hexamethonium. When ghrelin, applied intrathecally, was used to desensitize its receptors, the effect of intravenous CP464709 was blocked. CP464709 did not affect small intestine motility or the amplitudes of visceromotor reflexes caused by colorectal distension. It is concluded that activation of ghrelin receptors in the lumbo‐sacral spinal cord triggers co‐ordinated propulsive contractions that empty the colo‐rectum. The pathways through which these responses are generated pass out of the spinal cord via the pelvic nerves and cause propulsive contractions through activation of enteric neurons.

Functional Anatomy of Sympathetic Premotor Cell Groups in the Medulla

Pre- and postganglionic sympathetic neurons are organized into discrete functional channels, according to the target they supply. The activity patterns which they show differ between channels, implying that the CNS pathways driving them are not the same. Premotor neurons probably play a key role. This article focuses on what recent evidence tells us about the organization of premotor neurons which control specific sympathetic outflows. Cells that drive muscle vasoconstrictor (MVC), cutaneous vasoconstrictor (CVC), visceral vasoconstrictor (VVC) and renal sympathetic (RSN) outflows have been identified among the premotor neurons of the rostral ventrolateral medulla (RVLM). Other vasomotor, cardiac or adrenal drives are also represented in that cell group. Neurons driving sudomotor responses have been localized in the rostral ventromedial medulla. Evidence on the specific functions of other premotor cell groups is briefly discussed.

Hemorrhage induces c-fos immunoreactivity in spinally projecting neurons of cat subretrofacial nucleus.

Neuronal expression of c-fos product in the cat rostral ventrolateral medulla was studied by immunohistochemistry. Spinally projecting neurons of the subretrofacial (SRF) nucleus were pre-labeled by retrograde transport of fluorescent latex microspheres from the lumbar cord. In 3 animals which were bled by approximately 25% blood volume from a carotid cannula, 48-62% of SRF-spinal neurons expressed c-fos immunoreactivity. In 2 control animals, the corresponding values were 2% and 4%. The data show that bulbospinal neurons of presumed vasomotor function express c-fos in response to hemorrhage.

A quantitative approach to recording peristaltic activity from segments of rat small intestine in vivo

Abstract  We have developed methods that allow correlation of propulsive reflexes of the intestine with measurements of intraluminal pressure, fluid movement and spatio‐temporal maps of intestinal wall movements for the first time in vivo. A segment of jejunum was cannulated and set up in a Trendelenburg recording system while remaining connected to the vascular and nerve supply of the anaesthetized rat. The resting intraluminal pressure in intact intestine was 2–4 mmHg. Hydrostatic pressures of 2, 4, 8 and 16 mmHg were imposed. At a baseline pressure of 4 mmHg, propulsive waves generated pressures of 9 ± 1 mmHg, that progressed oral to anal at 2–5 mm s−1. Individual propulsive waves propelled 0.8 ± 0.4 mL of fluid. The frequency of propulsive waves increased with pressure, but peristaltic efficiency (mL per contraction) decreased with pressure increase between 4 and 16 mmHg. Atropine, as a bolus, transiently blocked peristalsis, but caused maintained block when infused. Hexamethonium blocked propulsive contractions. Inhibition of nitrergic transmission converted regular peristalsis to non‐propulsive contractions. These studies demonstrate the utility of an adapted Trendelenburg method for quantitative investigation of motility and pharmacology of enteric reflexes in vivo.

Intracellular responses of olfactory bulb granule cells to stimulating the horizontal diagonal band nucleus

The effects of centrifugal afferents on membrane potentials of identified granule cell layer using evoked field potential profiles, and trans-synaptic activation via antidromic stimulation of output cell axon collaterals. Intracellular recordings maintained for 4-30 min showed complex spontaneous spike discharges and allowed characterization of the cells input resistance, and on some occasions its morphology following intracellular injection of Lucifer Yellow. Stimulation in the nucleus of the horizontal limb of the diagonal band, but not surrounding regions, produced hyperpolarizing responses in 13 of 27 cells in the granule cell layer; four of these were morphologically identified as granule cells of two types, in five the responses had reversal potentials more negative than the resting potential, and six were identified as granule cells by monosynaptic activation from output axon collaterals. A different set of three cells in the granule cell layer responded with depolarization. The results are consistent with the inhibition of tonic activity of granule cells by the nucleus of the horizontal limb of the diagonal band, leading to disinhibition of mitral and tufted cells via dendrodendritic synapses of granule cells on mitral/tufted cell secondary dendrites.

Effect of stimulating the nucleus of the horizontal limb of the diagonal band on single unit activity in the olfactory bulb

The effects of centrifugal afferents on single unit discharge in the main olfactory bulb were studied in anaesthetized rats. Recording with extracellular micropipettes revealed spontaneous firing in all bulb layers. Units were located to different laminae using evoked field-potential profiles and histological verification. Output neurons were identified by antidromic response to stimulation of the lateral olfactory tract. Single- or brief multiple-pulse stimulation in the nucleus of the horizontal limb of the diagonal band, but not in adjacent regions, facilitated 17 out of 27 mitral cells with no effect on 10, but inhibited 21 out of 33 granule cell layer units with no effect on 12. Of 13 presumed tufted cells, six were facilitated and the rest unaffected. In contrast, stimulation of olfactory cortex inhibited mitral cells and facilitated most granule layer cells. The results are consistent with an inhibition of tonic granule cell discharge by the horizontal diagonal band nucleus, with resultant disinhibition of mitral cells via the dendrodendritic synapses of granule cells on mitral cell secondary dendrites.

Oral administration of a centrally acting ghrelin receptor agonist to conscious rats triggers defecation

Abstract  Agonists of ghrelin receptors that cross the blood–brain barrier, but not ghrelin itself, administered peripherally (intravenous or subcutaneous), cause defecation by acting on centres in the lumbo‐sacral spinal cord. It is not established whether orally administered ghrelin receptor agonists can have this action. We tested GSK894281 for its effectiveness at the ghrelin receptor and its ability to cross the blood–brain barrier. GSK894281 was effective at the human and rat ghrelin receptors at 1–10 nmol L−1, but was >1000‐fold less potent at the motilin receptor. It achieved a similar blood concentration by oral or intravenous administration. Oral bioavailability was 74% and brain : blood ratio at steady state was 0.7 : 1. GSK894281 administered orally (1–100 mg kg−1) caused a prompt, dose‐related production of faecal pellets; at 10 mg kg−1 faecal output was four times greater than after carrier. The output was the greatest in the first half hour and subsided over the next 90 min. At an oral dose of 10 mg kg−1, the compound was effective on eight successive days. Faecal output was, on average, increased threefold over control in the 2 h after administration on each of the 8 days. This dose also significantly increased food consumption. Rats showed no adverse behavioural effects to the drug on a single application, but at the end of a week of administration they avoided the gavaging pipette. Oral administration of ghrelin receptor agonists that enter the central nervous system could possibly be used to relieve acute cases of constipation or to clear the bowel for colonoscopy.

CRF-like immunoreactivity selectively labels preganglionic sudomotor neurons in cat

Immunohistochemical and neuronal tracing methods were used in cats to determine which type of postganglionic sympathetic neuron is innervated by preganglionic neurons which contain corticotrophin releasing factor-like immunoreactivity (CRF-LI). Preganglionic neurons with CRF-LI have their cell bodies at two restricted levels of the spinal cord and terminate in the stellate and lower lumbar ganglia. CRF-LI terminal baskets in stellate and lumbar ganglia surrounded cell bodies, 96-99% of which showed no tyrosine hydroxylase (TH)-LI (presumptive cholinergic neurons). Calcitonin gene-related peptide (CGRP)-LI was used to label the cholinergic ganglion cells which innervate sweat glands: 96-99% of those were confirmed as lacking TH-LI, while the remainder showed weak staining. Every one of over 6000 CRF-LI terminal baskets counted in 4 stellate and 6 lumbar ganglia was found to surround a cell body with CGRP-LI; conversely, 81-86% of the cell bodies showing CGRP-LI were surrounded by CRF-LI terminal baskets. In 3 cats, the retrograde tracer fluorogold was used to label postganglionic neurons projecting to the paw pads (a population which includes both cholinergic sudomotor neurons and noradrenergic vasoconstrictor neurons). Between 26 and 38% of the retrogradely labelled ganglion cells were surrounded by CRF-LI terminal baskets. We conclude that in cats, preganglionic sympathetic neurons with CRF-LI are sudomotor in function.

Motor patterns and propulsion in the rat intestine in vivo recorded by spatio‐temporal maps

Abstract  We have used spatio‐temporal maps derived from video images to investigate propagated contractions of the rat small intestine in vivo. The abdomen, including an exteriorized segment of jejunum, was housed in a humid chamber with a viewing window. Video records were converted to spatio‐temporal maps of jejunal diameter changes. Intraluminal pressure and fluid outflow were measured. Contractions occupied 3.8 ± 0.2 cm of intestine and propagated anally at 3.1 ± 0.2 mm s−1 when baseline pressure was 4 mmHg. Contractions at any one point lasted 8.7 ± 0.6 s. Contractions often occurred in clusters; within cluster frequencies were 2.28 ± 0.04 min−1. Pressure waves, with amplitudes greater than about 9 mmHg, expelled fluid when the baseline pressure was 4 mmHg. In the presence ofl‐NAME, circular muscle contractions occurred at a high frequency, but they were not propagated. We conclude that video recording methods give good spatio‐temporal resolution of intestinal movement when applied in vivo. They reveal neurally‐mediated propulsive contractions, similar to those previously recorded from intestinal segments in vitro. The propagated contractions had speeds of propagation that were slower and frequencies of occurrence that were less than speeds and frequencies of slow waves in the rat small intestine.