Antagonistic modulation of NPY/AgRP and POMC neurons in the arcuate nucleus by noradrenalin
- University of Cologne, Germany
- Excellence Cluster on Cellular Stress Responses in Aging Associated Diseases, Germany
- Max Planck Institute for Metabolism Research, Germany
- University Hospital Cologne, Germany
Figures
Figure 1
Noradrenalin differentially modulates functionally antagonistic NPY/AgRP and POMC neurons.
NA (10 µM) excited orexigenic NPY/AgRP neurons and inhibited anorexigenic POMC neurons. Original recordings (A) and averaged responses (B) of NPY/AgRP (n = 8) and POMC neurons (n = 8) during NA application. The mean response is expressed as change in action potential frequency. The boxplots show the absolute change in action potential frequency for both neuron populations. **p<0.01, Wilcoxon matched pairs signed ranks test.
Figure 2 with 1 supplement
Noradrenergic modulation of NPY/AgRP neurons is concentration dependent.
(A) Recording of a NPY/AgRP neuron demonstrating the effect of increasing NA concentrations. (B) and (C) Concentration - response relations showing the NA effect on action potential frequency (B) and membrane potential (C). The curves fit to a sigmoidal relation (Equation 1). NA had an EC50 of 1.9 µM (1.1–3.2 µM; n = 8) for the AP frequency and 1.5 µM (0.8–2.8 µM; n = 8) for the membrane potential, respectively. (D) and (E) The NA modulation of NPY/AgRP neurons is direct and not dependent on synaptic input. Original recording (D) and averaged effect on action potential frequency (E) showing that the NA effect on NPY/AgRP neurons is not changed when glutamatergic and GABAergic synaptic input is blocked. Experiments in (E) were performed consecutively with the same set of neurons (n = 8). Control 1 and control two refers to the different starting conditions, that is preincubation with or without synaptic blockers. ***p<0.001, one-way ANOVA with post hoc Tukey analysis.
Figure 2—figure supplement 1
Repeated applications of noradrenaline do not cause desensitization.
Original recording (A) of a NPY/AgRP neuron during two sequential applications of NA (10 µM) and averaged NA responses (B) on action potential frequency showing that there is no desensitization of the first application prior to the second treatment. n values are given in brackets. **p<0.01; one-way ANOVA with post hoc Tukey analysis.
Figure 3
Noradrenergic modulation of POMC neurons is concentration dependent.
(A) Recording of a POMC neuron demonstrating the effect of increasing NA concentrations. (B) and (C) Concentration - response relations showing the NA effect on membrane potential (B) and conductance density (C). The curves are fits to a sigmoidal relation (Equation 1). NA had an EC50 of 0.9 µM (0.6–1.5 µM; n = 8) for the membrane potential and 1.3 µM (1.0–1.9 µM; n = 8) for the conductance density, respectively. (D) and (E) The NA modulation of POMC neurons is direct and not dependent on synaptic input. Original recording (D) and averaged effect on conductance density (E) showing that the NA effect on POMC neurons is not changed when glutamatergic and GABAergic synaptic input is blocked. Experiments shown in (E) were performed consecutively with the same set of neurons (n = 4). Control 1 and control two refers to the different starting conditions, that is pre-incubation with or without synaptic blockers. **p<0.01; ***p<0.001; one-way ANOVA with post hoc Tukey analysis.
Figure 4 with 1 supplement
Cell type-specific expression of ARs.
(A) Log expression levels of adrenergic receptor genes in Pomc- and Npy-expressing cell populations. Deeper red colours indicate higher expression levels in the respective cell population. RPKM: reads per kilobase per million mapped reads. (B) Images from RNA in situ hybridizations against Adra1a (upper panels, green dots) and Adra2a (lower panels, green dots) in Npy- and Pomc- expressing (red) neurons. White arrowheads indicate doubly-labeled cells. Higher magnification indexes are representative of doubly-labeled cells. (C) Quantification of double positive cells for Adra1a and Adra2a in Npy- and Pomc- expressing neurons.
Figure 4—figure supplement 1
Images (A,B) and quantification (C) of RNA in situ hybridizations against Adrb1 (upper panels, green dots) and Adrb2 (lower panels, green dots) in Npy- and Pomc- expressing (red) neurons.
Higher magnification indexes are representative of doubly-labeled cells. Negative control: probe against E. coli gene DapB.
Figure 5
The excitatory noradrenergic effect on NPY/AgRP neurons is predominantly mediated by α1A-ARs.
(A) and (B) In 44% (4 out of 9) of the NPY/AgRP neurons the NA (10 µM) effect was completely blocked by the α1-AR antagonist prazosine (5 µM) and α2-AR antagonist yohimbine (5 µM) (A), while in 56% (5 out of 9) of the NPY/AgRP neurons the NA effect was markedly reduced (~60%) but not completely blocked (B). The series of experiments shown in (A) and (B) were each performed consecutively with the same set of neurons. n values are given in brackets. Control 1 and control two refers to the different starting conditions, that is pre-incubation with different AR antagonist. **p<0.01; ***p<0.001; one-way ANOVA with post hoc Tukey analysis.
Figure 6 with 1 supplement
NPY/AgRP neurons are excited by α1A-AR and inhibited by α2A-AR when α1A-AR are blocked.
(A) Rate histogram with its original recording showing that the α1A-AR agonist A61603 (1 µM) had similar excitatory effects on action potential frequency (B) as NA. (C) and (D) The selective α1A-AR antagonist WB4101 (100 nM) blocked the excitatory NA effect. In the presence of WB4101 NA had an inhibitory effect on the membrane potential, which was eliminated by the α2A-AR antagonist BRL44408 (10 µM). (E) The α1BD-AR blocker CEC (100 nM) did not change the NA effect on NPY/AgRP neurons. *p<0.05; **p<0.01; ***p<0.001; one-way ANOVA with posthoc Tukey analysis.
Figure 6—figure supplement 1
Repeated applications of noradrenaline do not cause desensitization.
Original recording (A) of a NPY/AgRP neuron during two sequential applications of A61603 (1 µM) and averaged A61603 responses (B) on action potential frequency showing that that there is no desensitization of the first application prior to the second treatment. n values are given in brackets. **p<0.01; one-way ANOVA with post hoc Tukey analysis.
Figure 7
The inhibitory noradrenalin effect on POMC neurons is mediated by α2A-AR.
(A) and (B) The α2A-antagonist BRL 44408 (10 µM) blocked the inhibitory NA (5 µM) effect (A), while the α2A-antagonist ARC239 (1 µM) did not affect the NA action (B). (C–E) Inhibitory NA effect (C). Blocking α2-AR by yohimbine revealed an excitatory NA effect (D), which was blocked by the α1-AR antagonist prazosine (E). The series of experiments shown in (A) and (B) were each performed consecutively with the same set of neurons. Control 1 and control two refers to the different starting conditions, that is preincubation with different AR antagonist. n values are given in brackets. *p<0.05; **p<0.01; ***p<0.001. (A) and (B) one-way ANOVA with post hoc Tukey analysis; (C–E) paired t-test.
Additional files
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Supplementary file 1
Significant gene expression differences between POMC- and NPY-expressing neurons.
FC, fold difference. p, p-value.
- https://doi.org/10.7554/eLife.25770.012
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Antagonistic modulation of NPY/AgRP and POMC neurons in the arcuate nucleus by noradrenalin
eLife 6:e25770.
https://doi.org/10.7554/eLife.25770
