The Expression of Galanin in the Parafacial Respiratory Group and its Effects on Respiration in Neonatal Rats.

The inhibitory peptide galanin is expressed within the retrotrapezoidal nucleus (RTN) - a key central chemoreceptor site that also contains the active expiratory oscillator. It was previously reported that microinjection of galanin into pre-Bötzinger complex - containing the inspiratory oscillator - exerts inhibitory effects on inspiratory motor output and respiratory rhythm. In neonatal rats, the present study aimed to investigate: (1) expression of galanin within the parafacial respiratory group (pFRG), which overlaps anatomically and functionally with the adult RTN, and; (2) effects of galanin on respiratory rhythm using the in vitro brainstem-spinal cord preparation. We showed that 14 ±â€¯2% of Phox2b-immunoreactive (ir) neurons in the parafacial region were also galanin-ir. Galanin peptide expression was confirmed within 3/9 CO2-sensitive, Phox2b-ir Pre-Inspiratory neurons (Pre-I) recorded in parafacial region. Bath application of galanin (0.1-0.2 µM): (1) decreased the duration of membrane depolarization in both Pre-I and inspiratory pFRG neurons, and; (2) decreased the number of C4 bursts that were associated with each burst in Pre-I neurons within the pFRG. In preparations showing episodic breathing at baseline, the respiratory patterning reverted to the 'normal' pattern of single, uniformly rhythmic C4 bursts (n = 10). In preparations with normal respiratory patterning at baseline, slowing of C4 rhythm (n = 7) resulted although rhythmic bursting in recorded Pre-I neurons remained unperturbed (n = 6). This study therefore demonstrates that galanin is expressed within the pFRG of neonatal rats, including neurons that are intrinsically chemosensitive. Overall the peptide has an inhibitory effect on inspiratory motor output, as previously shown in adults.

Brainstem galanin-synthesizing neurons are differentially activated by chemoreceptor stimuli and represent a subpopulation of respiratory neurons.

The ventrolateral medulla oblongata (VLM) of the brainstem contains neurochemically heterogeneous neurons that have a critical role in cardiovascular and respiratory regulation. Previous anatomical studies have shown the existence of galanin immunoreactivity in the medulla oblongata, but a detailed characterization is lacking. In this study, we demonstrate three populations of preprogalanin mRNA (PPG)-expressing neurons in the VLM of the adult, male Sprague-Dawley rat: a retrotrapezoid nucleus (RTN) group, a group in the rostral ventral respiratory group (rVRG), and a subpopulation of A1 neurons. PPG(+) neurons express tyrosine hydroxylase (TH) only in the A1 region of the VLM, where approximately 56% of PPG(+) neurons contain TH (79 ± 14; n = 4). PPG(+) neurons do not express vesicular acetylcholine transporter (vAChT) in the VLM (n = 3). However, 33% of PPG(+) neurons contain neurokinin-1 receptor (NK1R) in the rVRG (126 ± 12; n = 12), accounting for ∼28% of all NK1R(+) neurons in the region. Retrogradely transported cholera toxin B injected into the thoracic spinal cord (T1) revealed that bulbospinal PPG(+) neurons are present in the rVRG (n = 3; ∼26% of PPG(+) neurons). PPG(+) neurons in the RTN and locus coeruleus are selectively activated (Fos) following 2 hours of exposure to hypercapnia, but not by hypoxia. Neurons in the A1, nucleus of the solitary tract, and dorsomedial hypothalamus are activated by both chemoreceptor stimuli. The results suggest that PPG(+) neurons represent a population of brainstem neurons that play a critical and differential role in the chemoreflex responses to hypoxia and hypercapnia.

Differential regulation of the central neural cardiorespiratory system by metabotropic neurotransmitters.

Central neurons in the brainstem and spinal cord are essential for the maintenance of sympathetic tone, the integration of responses to the activation of reflexes and central commands, and the generation of an appropriate respiratory motor output. Here, we will discuss work that aims to understand the role that metabotropic neurotransmitter systems play in central cardiorespiratory mechanisms. It is well known that blockade of glutamatergic, gamma-aminobutyric acidergic and glycinergic pathways causes major or even complete disruption of cardiorespiratory systems, whereas antagonism of other neurotransmitter systems barely affects circulation or ventilation. Despite the lack of an 'all-or-none' role for metabotropic neurotransmitters, they are nevertheless significant in modulating the effects of central command and peripheral adaptive reflexes. Finally, we propose that a likely explanation for the plethora of neurotransmitters and their receptors on cardiorespiratory neurons is to enable differential regulation of outputs in response to reflex inputs, while at the same time maintaining a tonic level of sympathetic activity that supports those organs that significantly autoregulate their blood supply, such as the heart, brain, retina and kidney. Such an explanation of the data now available enables the generation of many new testable hypotheses.

Galanin is a selective marker of the retrotrapezoid nucleus in rats.

The rat retrotrapezoid nucleus (RTN) contains CO(2)-activated neurons that contribute to the central chemoreflex and to breathing automaticity. These neurons have two known markers, the transcription factor Phox2b and vesicular glutamate transporter 2 (VGLUT2). Noncatecholaminergic galanin-immunoreactive (ir) neurons within a region of the lower brainstem that seems identical to what is currently defined as the RTN have been previously described. Here we ask whether these galanin-expressing neurons are the same cells as the recently characterized CO(2)-sensitive neurons of the RTN. By using in situ hybridization, we found that pre-pro-galanin (PPGal) mRNA is expressed by an isolated cluster of neurons that is co-extensive with the RTN as defined by a population of strongly Phox2b-ir neurons devoid of tyrosine hydroxylase (Phox2b(+)/TH(-) neurons). This bilateral structure contains about 1,000 PPGal mRNA-positive neurons in the rat. The PPGal mRNA-positive neurons were Phox2b(+)/TH(-) and as susceptible to destruction by the toxin [Sar(9), Met (O(2))(11)]-substance P as the rest of the RTN Phox2b(+)/TH(-) cells of the RTN. CO(2)-activated neurons were recorded in the RTN of anesthetized rats and were labeled with biotinamide. Many of those cells (7/17, 41%, five rats) contained PPGal-mRNA. In conclusion, galanin mRNA is a very specific marker of the glutamatergic Phox2b(+)/TH(-) neurons of the RTN, but galanin mRNA identifies only half of these putative central respiratory chemoreceptors.

Metabotropic neurotransmission and integration of sympathetic nerve activity by the rostral ventrolateral medulla in the rat.

1. Cardiovascular sympathetic nerve activity at rest is grouped into waves, or bursts, that are generally, although not exclusively, related to the heart rate and to respiration. In addition, activity is also generated in response to central commands and to environmental stimuli. 2. Responsibility for the integration of all these different elements of sympathetic activity rests with pre-motoneurons in the rostral ventrolateral medulla oblongata. These pre-motoneurons are glutamatergic and spinally projecting where they form synapses with sympathetic preganglionic neurons. 3. Pre-motoneurons also contain and presumably release, neurotransmitters other than glutamate, including amines and neuropeptides that act on metabotropic receptors with long-term effects on cell function. 4. Similarly, in the rostral ventrolateral medulla oblongata the pre-motoneurons are mainly regulated by excitatory influences from glutamate and inhibitory influences from gamma-aminobutyric acid (GABA). Major focuses of recent studies are the interactions between non-glutamatergic and GABAergic systems and reflexes that regulate the activity of the sympathetic nervous system. 5. The results indicate that neurotransmitters acting at metabotropic receptors selectively affect different reflexes in the rostral ventrolateral medulla. It is suggested that this differential activation or attenuation of reflexes by different neurotransmitters is a mechanism by which the organism can fine-tune its responses to different homeostatic requirements.