Leonard F Arnolda

Bötzinger neurons project towards bulbospinal neurons in the rostral ventrolateral medulla of the rat.

Sympathetic nerve activity often fluctuates with the respiratory cycle, but the central neurons that impose this respiratory modulation have not been conclusively identified. In the present study, we used intracellular recording and dye‐filling to identify expiratory neurons in the Bötzinger complex. Our aim was to see if Bötzinger neurons project towards putative cardiovascular neurons in the rostral ventrolateral medulla. In the first series of experiments, histochemistry and immunohistochemistry were used to reveal the labelled Bötzinger neurons and neurons immunoreactive for tyrosine hydroxylase. Two out of four Bötzinger neurons had axon varicosities that were closely apposed to tyrosine hydroxylase‐immunoreactive neurons with cell bodies located within 0.6 mm caudal to the facial nucleus (three and five close appositions, respectively). In a second series of studies, rats were injected with cholera toxin B into the intermediolateral cell column of the spinal cord 4–7 days before the electrophysiological recording. Eight of the fourteen labelled Bötzinger neurons had a direct projection towards cholera toxin B‐labelled neurons in the rostral ventrolateral medulla. Close appositions were found on both somata and proximal dendrites (5 ± 2 close appositions/neuron, n=8). The present study supports the idea that a direct projection from Bötzinger neurons to presympathetic neurons in the rostral medulla plays a role in the respiratory modulation of sympathetic nerve activity. J. Comp. Neurol. 388:23–31, 1997.

Antisense to Thyrotropin Releasing Hormone Receptor Reduces Arterial Blood Pressure in Spontaneously Hypertensive Rats

We report in the present study the effect of intrathecal treatment with antisense oligonucleotides complementary to thyrotropin releasing hormone (TRH) receptor mRNA on the pressor response to intrathecal administration of TRH and on resting arterial blood pressure in Wistar-Kyoto (WKY) rats and spontaneously hypertensive rats (SHR). In 16-week-old male WKY rats, 18-base phosphodiester antisense or mismatch oligonucleotides to TRH receptor mRNA (100 micrograms per day) were injected intrathecally for 3 days. Twenty-four hours after the last injection, the magnitude of the pressor response to intrathecal TRH (10 micrograms) was significantly smaller in the antisense-treated group (n = 7) compared with mismatch-treated controls (n = 7) (change in mean arterial pressure, +20.3 +/- 3.0 versus +32.6 +/- 2.5 mm Hg, P < .01). No differences were observed in the pressor responses to injection of N-methyl-D-aspartic acid. Resting arterial blood pressure was unaffected by antisense treatment in WKY rats. In separate experiments, 16-week-old male SHR were treated with antisense (n = 7) or mismatch (n = 6) oligonucleotides for 3 days. Mean resting arterial blood pressure was significantly reduced by treatment with antisense oligonucleotides (from 157 +/- 4.8 to 119 +/- 8.8 mm Hg, P < .01), but no significant changes were observed in mismatch-treated animals. Our results suggest that the expression of TRH receptors in spinal sympathetic preganglionic neurons can be selectively reduced by intrathecal treatment with antisense oligonucleotides and that TRH projections to sympathetic preganglionic neurons play an important role in the elevation of arterial blood pressure in SHR.

AMPA/kainate receptors mediate sympathetic chemoreceptor reflex in the rostral ventrolateral medulla

Previous studies have reported that information from carotid chemoreceptors activates sympathetic premotor neurons in the rostral ventrolateral medulla (RVLM) exclusively via N-methyl-D-aspartic acid (NMDA) receptors. In this study, we examined the possible involvement of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainate receptors in the RVLM on sympathetic chemoreceptor reflex in pentobarbitone anaesthetised, vagotomised and artificially ventilated rats. Carotid chemoreceptor stimulation with brief N2 inhalation increased splanchnic sympathetic nerve activity and arterial pressure in animals that had received an intravenous injection of the non-competitive NMDA receptor blocker, MK-801 (2 mg/kg). RVLM sympathetic premotor neurons could also be activated by brief hypoxia in the presence of MK-801. However, microinjection of 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, a selective AMPA/kainate receptor antagonist, 2 mM, 100 nl) into the RVLM after intravenous MK-801 abolished the hypoxia evoked sympathoexcitatory response. These results demonstrate that AMPA/kainate receptors in the RVLM are involved in the chemoreceptor reflex pathway.

CENTRAL NEURONS AND NEUROTRANSMITTERS IN THE CONTROL OF BLOOD PRESSURE

1. In this paper we review recent work from our laboratory on two major pathways important in the central control of blood pressure.

Low-dose diuretic and/or dietary sodium restriction when blood pressure is resistant to ACE inhibitor.

AIMnTo compare the efficacy of indapamide (1.25 mg daily) and low-salt diet (<100 mmol/day) separately and in combination in essential hypertensive patients with inadequate BP response to perindopril.nnnDESIGN AND METHODSnRandomized double-blind, double-dummy, crossover design. The randomized treatments were indapamide 1.25 mg daily, sodium chloride 80 mmol daily, the combination of indapamide and sodium chloride and placebo. All patients received perindopril 4 mg daily and maintained a low-sodium diet.nnnRESULTSn19 patients entered and 17 completed the study. Prior to randomization, average clinic sitting blood pressure was 162/101 mm Hg and average 24-h urine sodium excretion was 157 mmol/day. Compared to the phase in which patients received perindopril with sodium repletion, clinic and ambulatory BPs were significantly reduced (p<0.01) in all the other phases. Indapamide had a greater effect on BP than dietary sodium restriction, and in combination their effects were additive. The effect of indapamide on ambulatory BP persisted throughout 24 h, but the effect of the low-salt diet was predominantly observed during waking hours.nnnCONCLUSIONSnIn hypertensives with BP resistant to the angiotensin converting enzyme (ACE) inhibitor perindopril, the diuretic indapamide had greater additional efficacy and longer duration of action than dietary sodium restriction. In combination they had additive effects on BP.

Thyrotropin-releasing hormone immunoreactive boutons form close appositions with medullary expiratory neurons in the rat

The aim of the present study was to assess the size of the input from TRH immunoreactive varicosities to medullary respiratory neurons in the Bötzinger complex and caudal ventral respiratory group. Neurobiotin was intracellularly injected into seven neurons in the Bötzinger complex, between 0.4 and 0.9 mm caudal to the facial nucleus. Five of the seven Bötzinger neurons had extensive local axonal projections, with bouton-like varicosities clustered predominantly between their somata and the nucleus ambiguus. Seven neurons in the caudal ventral respiratory group, located between 1.6 and 2.4 mm caudal to the facial nucleus, were also labelled. All but one caudal respiratory neurons had no, or very few, medullary collaterals. TRH immunoreactive fibres were seen in many medullary nuclei, including the ventral reticular formation. Bötzinger neurons were closely apposed by an average of 29 +/- 8 TRH immunoreactive boutons/neuron (mean +/- S.D., n = 7). In contrast, caudal ventral respiratory group neurons were apposed by only 5 +/- 3 TRH immunoreactive boutons/neuron (n = 7). Bötzinger neurons form many intramedullary and bulbospinal inhibitory connections with premotoneurons and motoneurons that are important in the timing, amplitude and shape, of respiratory activity. Our findings suggest a role for endogenous TRH-containing neurons in modulating the activity of inhibitory Bötzinger neurons and neurons in the caudal ventral respiratory group. The significance of the apparent difference in size of this input remains to be determined.

Role of spinal GABA receptors in depressor responses to chemical stimulation of the A5 area in normal and hypertensive rats

Chemical stimulation of neurons in the pontine A5 area by microinjection of L-glutamate lowers arterial blood pressure. The mechanism of this A5 depressor response is not well-established. Here, we examine the involvement of spinal cord gamma-aminobutyric acid (GABA) receptors in this depressor response in normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR). Experiments were conducted in male WKY and age-matched SHR anaesthetised with sodium pentobarbitone and chloral hydrate. An intrathecal catheter was implanted with the tip located between T9 and L2. Three days later, rats were re-anaesthetised and 10 nl of 40 mM L-glutamate was injected into the A5 area before, during and after, blockade of spinal cord GABA-A receptors by intrathecal injection of bicuculline methiodide (1 mM in 10 microliters phosphate-buffered saline). Injection of L-glutamate (10, 20, 40, 80 mM in 10 nl) produced depressor responses that were similar in WKY (n = 6) and SHR (n = 6). Intrathecal injection of bicuculline elicited a pressor response that was greater in SHR (n = 7, 28.5 +/- 7.6% increase in mean arterial pressure) than WKY (n = 11, 11.6 +/- 3.6%, p < 0.05). After bicuculline, the depressor response to injection of L-glutamate into the A5 area was eliminated in both WKY (n = 7) and SHR (n = 6). Intrathecal injection of vehicle had no effect on either resting arterial blood pressure or the depressor response to A5 stimulation. Basal blood pressure and control responses to A5 stimulation were fully restored by around 90 min after bicuculline injection in each animal. In separate groups of rats, intrathecal injection of muscimol elicited depressor responses that were greater in SHR (n = 6, -32.0 +/- 6.2%) than WKY (n = 6, -17.3 +/- 1.5%, p < 0.05). Our results suggest that the A5 depressor response is due to a projection from the A5 area to the spinal cord. This projection acts directly, or through a spinal interneuron and uses GABA as a neurotransmitter. Furthermore, our results indicate a hyper-responsiveness to GABA-A receptor stimulation in SHR since intrathecal bicuculline elicited much greater increases and intrathecal muscimol elicited much greater decreases, in blood pressure in SHR than in WKY. Finally, it seems likely that the A5-spinal depressor pathway is less effective in SHR than WKY under physiological conditions since chemical stimulation of the A5 area with L-glutamate produced a comparable depressor response in both strains.

Substance p and serotonergic inputs to sympathetic preganglionic neurons

Sympathetic preganglionic neurons are the final central links in the sympathetic pathways that control the heart and blood vessels. The neurotransmitters present in the supraspinal pathways that control the activity of sympathetic preganglionic neurons include amino acids, amines and peptides. In this paper we discuss evidence that suggests a role for serotonin and substance P in these pathways. Both of these neurotransmitters are present in bulbospinal neurons. Our results suggest that they have an important physiological role in the central regulation of blood pressure.

Tachycardia after glutamate injection in rat spinal cord is not blocked by kynurenate or mimicked by metabotropic agonists.

1. We have used microinjections of glutamate, an ionotropic excitatory amino acid receptor antagonist (kynurenate) and selective ionotropic (NMDA and kainate) and metabotropic (lS‐3R‐ACPD, trans‐ACPD and L‐AP4) receptor agonists in the thoracic IML of the rat to define the receptors mediating the tachycardia produced by excitatory amino acid agonists.