Transgenic rodents as dynamic models for the study of respiratory rhythm generation and modulation: a scoping review and a bibliometric analysis.

The pre-Bötzinger complex, situated in the ventrolateral medulla, serves as the central generator for the inspiratory phase of the respiratory rhythm. Evidence strongly supports its pivotal role in generating, and, in conjunction with the post-inspiratory complex and the lateral parafacial nucleus, in shaping the respiratory rhythm. While there remains an ongoing debate concerning the mechanisms underlying these nuclei's ability to generate and modulate breathing, transgenic rodent models have significantly contributed to our understanding of these processes. However, there is a significant knowledge gap regarding the spectrum of transgenic rodent lines developed for studying respiratory rhythm, and the methodologies employed in these models. In this study, we conducted a scoping review to identify commonly used transgenic rodent lines and techniques for studying respiratory rhythm generation and modulation. Following PRISMA guidelines, we identified relevant papers in PubMed and EBSCO on 29 March 2023, and transgenic lines in Mouse Genome Informatics and the International Mouse Phenotyping Consortium. With strict inclusion and exclusion criteria, we identified 80 publications spanning 1997-2022 using 107 rodent lines. Our findings revealed 30 lines focusing on rhythm generation, 61 on modulation, and 16 on both. The primary in vivo method was whole-body plethysmography. The main in vitro method was hypoglossal/phrenic nerve recordings using the en bloc preparation. Additionally, we identified 119 transgenic lines with the potential for investigating the intricate mechanisms underlying respiratory rhythm. Through this review, we provide insights needed to design more effective experiments with transgenic animals to unravel the mechanisms governing respiratory rhythm. The identified transgenic rodent lines and methodological approaches compile current knowledge and guide future research towards filling knowledge gaps in respiratory rhythm generation and modulation.

Perinatal inflammation and gestational intermittent hypoxia disturbs respiratory rhythm generation and long-term facilitation in vitro: Partial protection by acute minocycline.

Perinatal inflammation triggers breathing disturbances early in life and affects the respiratory adaptations to challenging conditions, including the generation of amplitude long-term facilitation (LTF) by acute intermittent hypoxia (AIH). Some of these effects can be avoided by anti-inflammatory treatments like minocycline. Since little is known about the effects of perinatal inflammation on the inspiratory rhythm generator, located in the preBötzinger complex (preBötC), we tested the impact of acute lipopolysaccharide (LPS) systemic administration (sLPS), as well as gestational LPS (gLPS) and gestational chronic IH (gCIH), on respiratory rhythm generation and its long-term response to AIH in a brainstem slice preparation from neonatal mice. We also evaluated whether acute minocycline administration could influence these effects. We found that perinatal inflammation induced by sLPS or gLPS, as well as gCIH, modulate the frequency, signal-to-noise ratio and/or amplitude (and their regularity) of the respiratory rhythm recorded from the preBötC in the brainstem slice. Moreover, all these perinatal conditions inhibited frequency LTF and amplitude long-term depression (LTD); gCIH even induced frequency LTD of the respiratory rhythm after AIH. Some of these alterations were not observed in slices pre-treated in vitro with minocycline, when compared with slices obtained from naïve pups, suggesting that ongoing inflammatory conditions affect respiratory rhythm generation and its plasticity. Thus, it is likely that alterations in the inspiratory rhythm generator and its adaptive responses could contribute to the respiratory disturbances observed in neonates that suffered from perinatal inflammatory challenges.

Configuration and dynamics of dominant inspiratory multineuronal activity patterns during eupnea and gasping generation in vitro.

The pre-Bötzinger complex (preBötC), located within the ventral respiratory column, produces inspiratory bursts in varying degrees of synchronization/amplitude. This wide range of population burst patterns reflects the flexibility of the preBötC neurons, which is expressed in variations in the onset/offset times of their activations and their activity during the population bursts, with respiratory neurons exhibiting a large cycle-to-cycle timing jitter both at the population activity onset and at the population activity peak, suggesting that respiratory neurons are stochastically activated before and during the inspiratory bursts. However, it is still unknown whether this stochasticity is maintained while evaluating the coactivity of respiratory neuronal ensembles. Moreover, the preBötC topology also remains unknown. In this study, by simultaneously recording tens of preBötC neurons and using coactivation analysis during the inspiratory periods, we found that the preBötC has a scale-free configuration (mixture of not many highly connected nodes, hubs, with abundant poorly connected elements) exhibiting the rich-club phenomenon (hubs more likely interconnected with each other). PreBötC neurons also produce multineuronal activity patterns (MAPs) that are highly stable and change during the hypoxia-induced reconfiguration. Moreover, preBötC contains a coactivating core network shared by all its MAPs. Finally, we found a distinctive pattern of sequential coactivation of core network neurons at the beginning of the inspiratory periods, indicating that, when evaluated at the multicellular level, the coactivation of respiratory neurons seems not to be stochastic.NEW & NOTEWORTHY By means of multielectrode recordings of preBötC neurons, we evaluated their configuration in normoxia and hypoxia, finding that the preBötC exhibits a scale-free configuration with a rich-club phenomenon. preBötC neurons produce multineuronal activity patterns that are highly stable but change during hypoxia. The preBötC contains a coactivating core network that exhibit a distinctive pattern of coactivation at the beginning of inspirations. These results reveal some network basis of inspiratory rhythm generation and its reconfiguration during hypoxia.

Local cortical activity of distant brain areas can phase-lock to the olfactory bulb's respiratory rhythm in the freely behaving rat.

An important unresolved question about neural processing is the mechanism by which distant brain areas coordinate their activities and relate their local processing to global neural events. A potential candidate for the local-global integration are slow rhythms such as respiration. In this study, we asked if there are modulations of local cortical processing that are phase-locked to (peripheral) sensory-motor exploratory rhythms. We studied rats on an elevated platform where they would spontaneously display exploratory and rest behaviors. Concurrent with behavior, we monitored whisking through electromyography and the respiratory rhythm from the olfactory bulb (OB) local field potential (LFP). We also recorded LFPs from dorsal hippocampus, primary motor cortex, primary somatosensory cortex, and primary visual cortex. We defined exploration as simultaneous whisking and sniffing above 5 Hz and found that this activity peaked at ~8 Hz. We considered rest as the absence of whisking and sniffing, and in this case, respiration occurred at ~3 Hz. We found a consistent shift across all areas toward these rhythm peaks accompanying behavioral changes. We also found, across areas, that LFP gamma (70-100 Hz) amplitude could phase-lock to the animal's OB respiratory rhythm, a finding indicative of respiration-locked changes in local processing. In a subset of animals, we also recorded the hippocampal theta activity and found that occurred at frequencies overlapped with respiration but was not spectrally coherent with it, suggesting a different oscillator. Our results are consistent with the notion of respiration as a binder or integrator of activity between brain regions.

Differential Expression of Ion Channels in Adult and Neonatal Rat Ventral Respiratory Column.

In mammals, the neural control of breathing is attributed to circuits distributed along the ventral respiratory column (VRC) in the ventrolateral medulla. The VRC contains the kernel for generation of the inspiratory phase of respiratory rhythm and nuclei involved in central chemoreception. During development, the respiratory rhythm, as well as central chemosensitivity, adjusts to meet the changing physiological requirements associated with increased body weight and size. Gene expression in VRC ontogeny is well characterized. However, little is known about gene expression in the VRC during postnatal development. Here, we sought to characterize the changes in gene expression that occur in the VRC of the adult rat (5-6 months of age) in comparison with the VRC of neonate rat (1-4 days old). We isolated total RNA from VRC tissue punches collected from thick transversal slices. We hybridized cDNA to a 5000-oligonucleotide rat microarray. We found that 218 genes (4.4%) of the 5000 genes in the microarray changed their expression in adult VRC with respect to that from neonate. To further analyze the modified expression of specific genes, we quantified the differential expression of 84 genes of neuronal ion channels using a quantitative RT-PCR array. This analysis confirmed the overexpression of 68 genes and the underexpression of 14 genes in the VRC from adult compared with that from neonate. Our findings may help to explain the functional changes in respiratory rhythm and chemosensitivity occurring throughout life.

The Onset of the Fetal Respiratory Rhythm: An Emergent Property Triggered by Chemosensory Drive?

The mechanisms responsible for the onset of respiratory activity during fetal life are unknown. The onset of respiratory rhythm may be a consequence of the genetic program of each of the constituents of the respiratory network, so they start to interact and generate respiratory cycles when reaching a certain degree of maturation. Alternatively, generation of cycles might require the contribution of recently formed sensory inputs that will trigger oscillatory activity in the nascent respiratory neural network. If this hypothesis is true, then sensory input to the respiratory generator must be already formed and become functional before the onset of fetal respiration. In this review, we evaluate the timing of the onset of the respiratory rhythm in comparison to the appearance of receptors, neurotransmitter machinery, and afferent projections provided by two central chemoreceptive nuclei, the raphe and locus coeruleus nuclei.

Role of Astrocytes in Central Respiratory Chemoreception.

Astrocytes perform various homeostatic functions in the nervous system beyond that of a supportive or metabolic role for neurons. A growing body of evidence indicates that astrocytes are crucial for central respiratory chemoreception. This review presents a classical overview of respiratory central chemoreception and the new evidence for astrocytes as brainstem sensors in the respiratory response to hypercapnia. We review properties of astrocytes for chemosensory function and for modulation of the respiratory network. We propose that astrocytes not only mediate between CO2/H+ levels and motor responses, but they also allow for two emergent functions: (1) Amplifying the responses of intrinsic chemosensitive neurons through feedforward signaling via gliotransmitters and; (2) Recruiting non-intrinsically chemosensitive cells thanks to volume spreading of signals (calcium waves and gliotransmitters) to regions distant from the CO2/H+ sensitive domains. Thus, astrocytes may both increase the intensity of the neuron responses at the chemosensitive sites and recruit of a greater number of respiratory neurons to participate in the response to hypercapnia.

Control of respiration in fish, amphibians and reptiles

Fish and amphibians utilise a suction/force pump to ventilate gills or lungs, with the respiratory muscles innervated by cranial nerves, while reptiles have a thoracic, aspiratory pump innervated by spinal nerves. However, fish can recruit a hypobranchial pump for active jaw occlusion during hypoxia, using feeding muscles innervated by anterior spinal nerves. This same pump is used to ventilate the air-breathing organ in air-breathing fishes. Some reptiles retain a buccal force pump for use during hypoxia or exercise. All vertebrates have respiratory rhythm generators (RRG) located in the brainstem. In cyclostomes and possibly jawed fishes, this may comprise elements of the trigeminal nucleus, though in the latter group RRG neurons have been located in the reticular formation. In air-breathing fishes and amphibians, there may be separate RRG for gill and lung ventilation. There is some evidence for multiple RRG in reptiles. Both amphibians and reptiles show episodic breathing patterns that may be centrally generated, though they do respond to changes in oxygen supply. Fish and larval amphibians have chemoreceptors sensitive to oxygen partial pressure located on the gills. Hypoxia induces increased ventilation and a reflex bradycardia and may trigger aquatic surface respiration or air-breathing, though these latter activities also respond to behavioural cues. Adult amphibians and reptiles have peripheral chemoreceptors located on the carotid arteries and central chemoreceptors sensitive to blood carbon dioxide levels. Lung perfusion may be regulated by cardiac shunting and lung ventilation stimulates lung stretch receptors.