Neuronal Mediators of the Frequency Increase Evoked by Activation of P2Y <sub>1</sub> Receptors in the preBötzinger Complex Inspiratory Rhythm Generating Network in Vitro
Bibliographic record
Abstract
The ventilatory response to acute hypoxia (HVR) comprises an initial, carotid body‐mediated increase in breathing followed by a centrally‐mediated secondary respiratory depression when ventilation falls to a new steady state. Recent evidence suggests that ATP is released from astrocytes in the preBötzinger Complex (preBötC; site of inspiratory rhythm generation) during the secondary phase where it acts via P2Y 1 receptors (Rs) to excite preBötC neurons, increase breathing frequency and attenuate the secondary respiratory depression. The preBötC network comprises a heterogenous population of excitatory and inhibitory inspiratory neurons that can be further classified based on expression of transcription factors, receptors and transmitters, discharge pattern and membrane properties. An important goal is to establish the neuronal phenotype(s) that underlie the P2Y 1 R frequency increase. Perhaps surprisingly, preBötC frequency increases in response to optogenetic stimulation of excitatory OR inhibitory preBötC neurons. Thus, the objective of this study was to determine if the P2Y 1 R‐mediated frequency increase results from activation of excitatory (glutamatergic) or inhibitory (GABA/glycinergic) preBötC inspiratory neurons. Analysis of P2Y 1 R and somatostatin (SST) immunolabeling in the medulla of neonatal vGLUT2 TdTomato and GLYT2 EGFP reporter mice showed that P2Y 1 R immunolabeled neurons are concentrated in a discrete region of the ventral respiratory column (VRC) that features strong SST immunolabeling (a preBötC marker). P2Y 1 R+ neurons were sparse in all other regions of the VRC. In addition, 95% of P2Y 1 R+ neurons were glutamatergic (VGLUT2+), but <10% of glutamatergic neurons were P2Y 1 R+. None of the GLYT2+ (glycinergic) preBötC neurons were P2Y 1 R+. Whole‐cell analyses of neurons in rhythmic slices from vGLUT2 mice showed that all VGLUT2+ inspiratory preBötC neurons excited by the P2Y 1 R agonist MRS2365 (100 µM) were glutamatergic, but only1/3 of VGLUT2+ neurons were P2Y 1 R sensitive. In contrast, calcium imaging of rhythmic slices identified subpopulations (4/16) of glutamatergic and non glutamatergic inspiratory neurons that in TTX were P2Y 1 R sensitive. Finally, we demonstrated that the frequency increase evoked by MRS2365 (100 µM) in the preBötC was unaffected by block of inhibitory synaptic transmission via GABA A (bicuculline, 20 µM) and glycine (strychnine, 20 µM) receptor antagonists. In summary, we found that P2Y 1 R‐expressing neurons are concentrated in the preBötC and are primarily glutamatergic. P2Y 1 R‐sensitive neurons are primarily glutamatergic, but a subpopulation is nonglutamatergic. These data, combined with the observation that the P2Y 1 R‐evoked excitation of the preBötC does not depend on synaptic inhibition, suggest that the P2Y 1 R‐mediated excitation of the preBötC is primarily achieved through activation of excitatory, glutamatergic neurons.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".