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Perineuronal net formation during the critical period for neuronal maturation in the hypothalamic arcuate nucleus


In leptin-deficient ob/ob mice, obesity and diabetes are associated with abnormal development of neurocircuits in the hypothalamic arcuate nucleus (ARC)1, a critical brain area for energy and glucose homoeostasis2,3. Because this developmental defect can be remedied by systemic leptin administration, but only if given before postnatal day 28, a critical period for leptin-dependent development of ARC neurocircuits has been proposed4. In other brain areas, critical-period closure coincides with the appearance of perineuronal nets (PNNs), extracellular matrix specializations that restrict the plasticity of neurons that they enmesh5. Here we report that in humans and rodents, subsets of neurons in the mediobasal aspect of the ARC are enmeshed in PNN-like structures. In mice, these neurons are densely packed into a continuous ring that encircles the junction of the ARC and median eminence, which facilitates exposure of ARC neurons to the circulation. Most of the enmeshed neurons are both γ-aminobutyric acid-ergic and leptin-receptor positive, including a majority of Agouti-related-peptide neurons. Postnatal formation of the PNN-like structures coincides precisely with closure of the critical period for maturation of Agouti-related-peptide neurons and is dependent on input from circulating leptin, because postnatal ob/ob mice have reduced ARC PNN-like material that is restored by leptin administration during the critical period. We conclude that neurons crucial to metabolic homoeostasis are enmeshed in PNN-like structures and organized into a densely packed cluster situated circumferentially at the ARC–median eminence junction, where metabolically relevant humoral signals are sensed.

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Fig. 1: WFA labelling in the ventromedial ARC forms a ‘collar’ around the median eminence.
Fig. 2: PNNs enmesh GABAergic, LepRb+, AgRP/NPY neurons in the ARC.
Fig. 3: PNN formation in the ARC occurs during the lactation and periweaning period, corresponding to the maturation of AgRP neurons.
Fig. 4: Leptin-deficient ob/ob mice have impaired PNN formation during postnatal development that can be rescued by leptin administration during the critical period.

Data availability

The data that support the findings of this study are available from the corresponding authors upon request. Additional detailed information on experimental design and reagents is available in the Reporting Summary.


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The authors are grateful to the original providers of the transgenic mouse lines used in this work: N. Tamamaki (GAD67-GFP), M. Myers (LepRb-Cre), H. Zeng (Ai14), B. Lowell (NPY-GFP) and M. Low (POMC-GFP). This work was supported by the US National Institutes of Health National Institute of Diabetes and Digestive and Kidney Diseases (grant nos. DK108596 (Z.M.), DK114474 (J.M.S.), DK083042 (M.W.S.), DK090320 (M.W.S.) and DK101997 (M.W.S.)), the National Institute of Neurological Disorders and Stroke Neurosurgeon Research Career Development Program K Award (Z.M.), the American Diabetes Association (grant no. 7–11-BS-179 (L.Z.)), the Russell Berrie Foundation (R.H.) and the Barrow Neurological Foundation (grant no. 18-0025-30-05 (Z.M.)).

Author information

Authors and Affiliations



Z.M., K.M.A. and M.W.S. conceived and designed the study; Z.M., K.M.A., E.C., V.H.P., J.M.S., J.M.B., R.H., M.E.M., H.T.N., J.M.G.V., L.M.Z. and M.W.S. acquired, analysed and interpreted the data; Z.M. and M.W.S. drafted and revised the manuscript. All authors approved the final version of the manuscript.

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Correspondence to Zaman Mirzadeh or Michael W. Schwartz.

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Supplementary information

Supplementary Information

Supplementary Figures. 1–9

Reporting Summary

Supplementary Video 1

Imaris three-dimensional (3D) surface rendering of an isolated PNN-enmeshed (red) GAD67-GFP+ ARC neuron (green). This rendering was performed on a 63× image taken from a coronal section, through the ARC of a GAD67-GFP mouse, stained with WFA (red) and GFP antibody (green). This 3D image is representative of renderings performed on five other GAD67-GFP+ ARC neurons

Supplementary Video 2

Imaris 3D surface rendering of an isolated PNN-enmeshed (red) GAD67-GFP+ V1 interneuron (green). This rendering was performed on a 63× image taken from a coronal section, through V1 of a GAD67-GFP mouse, stained with WFA (red) and GFP antibody (green). This 3D image is representative of renderings performed on three other GAD67-GFP+ V1 neurons.

Supplementary Video 3

Imaris 3D surface rendering of an isolated PNN-enmeshed (red) Npy-GFP+ ARC neuron (green). This rendering was performed on a 63× image taken from a coronal section, through the ARC of a Npy-GFP mouse, stained with WFA (red) and GFP antibody (green). This 3D image is representative of renderings performed on five other NPY-GFP+ neurons.

Supplementary Video 4

Imaris 3D surface rendering of an isolated PNN-enmeshed (red) NPY+ human ARC neuron (green). This rendering was performed on a 63× image taken from a coronal section, through the ARC of a human brain, stained with WFA (red) and NPY antibody (green). This 3D image is representative of renderings performed on four other NPY+ human ARC neurons.

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Mirzadeh, Z., Alonge, K.M., Cabrales, E. et al. Perineuronal net formation during the critical period for neuronal maturation in the hypothalamic arcuate nucleus. Nat Metab 1, 212–221 (2019).

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