05 Jan 2024

Neurons intertwine with small blood vessels in the brain, but how do the two communicate? With elaborate three-dimensional reconstructions of electron microscopy images, scientists led by Jie-Min Jia, Westlake University in Hangzhou, China, found that when axons snuggle up to smooth muscle cells in arterioles, they connect via structures that seem awfully similar to synapses. In the January 2 Nature Neuroscience, the scientists reported that glutamate travels across these “neuromuscular junctions” to bind receptors on the smooth muscle.

The cells then relaxed, allowing the vessels to dilate and more blood to flow. Knocking out the glutamate receptor on these smooth muscle cells weakened this neurovascular coupling. Knowing that glutamate directly controls vasodilation could give researchers new ways to manipulate neurovascular interactions. Reduced neurovascular coupling has been linked to cognitive decline and Alzheimer’s disease.

“This work provides further support to the idea that neurons within the brain directly contact arteriole smooth muscle cells. It sheds new light on the intricacies of how these interactions regulate cerebral perfusion during brain activity,” Costantino Iadecola, Weill Cornell Medical College, New York, told Alzforum. Shahram Oveisgharan, Rush University Medical Center in Chicago, agreed. “The findings are very intriguing as [they] extend possible mechanisms underlying neurovascular coupling,” he wrote (comment below).

Researchers previously proposed that some perivascular nerves contact arteriole smooth muscle cells (aSMCs), suggesting the neurons could directly, via neurotransmitters, signal vessels to relax (Krimer et al., 1998; reviewed by Sandor, 1999). This concept was hotly debated. The general consensus is that vasodilation results from nitric oxide, carbon dioxide, adenosine, and other molecules produced as a result of neural activity, often by astrocytes, not from direct neurotransmission (Iadecola, 2017).

To see neuron-vessel interactions up close, first author Dongdong Zhang took 3,000 serial electron microscopy images of a block of mouse cerebral cortex tissue and created a three-dimensional reconstruction of the cellular interactions surrounding arterioles. Astrocyte end feet usually coat arterioles, but Zhang found that there were gaps. What's more, dendrites and axons threaded in to those gaps to touch aSMCs, forming a type of neuromuscular junction (image and video below). On average, each aSMC had two or three such junctions where it was touched by a dendrite.

Neurovascular Junctions.  In an arteriole (red) of the mouse brain, astrocyte end feet (cyan) leave gaps for axons (green) to contact the arteriole. Click for a video of the reconstructed arteriole. [Courtesy of Zhang et al., Nature Neuroscience, 2024.]

What was happening at the junctions? Eighty percent of them contained small vesicles, suggesting that they may carry neurotransmitters. Indeed, axons expressing the glutamate transporter vGluT1 innervated aSMCs. Bulk RNA-Seq of isolated smooth muscle cells indicated they express a range of neurotransmitter receptors, including those for glutamate. Gene expression signatures confirmed no neuron or endothelial cell contamination. Puncta of the GluN1 subunit of the NMDA glutamate receptor clustered on the outer cell membrane of the smooth muscle cells, where it might directly interact with neurons.

Zhang saw the same distribution of GluN1 on the outside of aSMCs in brain tissue from a macaque and a person, suggesting glutamatergic signaling at neurovascular junctions might be universal in mammals. “Future human studies are needed to confirm the expression of glutamate receptors in smooth muscle cells of cerebral small vessels,” noted Oveisgharan.

Notably, in cultured mouse aSMCs, almost all GluN1 co-localized with the postsynaptic scaffold protein PSD95, which is expressed in vascular smooth muscle cells (Joseph et al., 2011; Moore et al., 2014). In co-cultures of aSMCs and neurons, GluN1-expressing muscle cells also cozied up to neurons expressing vGluT1. The authors take this to mean that these neurovascular junctions might behave like synapses.

To test this, Zhang first added glutamate to cultured aSMCs. This evoked an electrical current within the cells, enabling them to relax or contract, depending on their physiological context.

In living mice, optogenetically activating glutamatergic neurons surrounding arterioles dilated the vessels (image below). On the other hand, selectively knocking out GluN1 in aSMCs disrupted neurovascular communication. It halved vasodilation induced by stimulation of glutamatergic neurons and dampened cerebral blood flow after tickling the animals’ whiskers, a standard test of neurovascular coupling.

Magic Opener. Stimulating a neuron (green) next to an arteriole (red) dilates the vessel. [Courtesy of Zhang et al., Nature Neuroscience, 2024.]

Do these glutamate-driven neurovascular couplings relate to Alzheimer’s disease? Some evidence exists to suggest that Aβ restricts cerebral blood flow, and Iadecola noted that tauopathy mice have only half the surge in blood flow after their whiskers get tickled than do wild-type mice (Jun 2019 news; Aug 2020 news). He found that this deficit in neurovascular coupling stems from soluble phospho-tau binding to PSD95/NMDA complexes in postsynapses, preventing neuronal nitric oxide synthase (nNOS) from binding the complex and making vasodilatory nitric oxide. “Since glutamatergic nNOS neurons are known to contact cerebral arterioles, tau might also lead to neurovascular uncoupling through the GluN1/NMDA mechanism suggested by Zhang and colleagues,” he said.—Chelsea Weidman Burke

Comments

1. • Shahram Oveisgharan
     Rush University Medical Center
   • Posted: 05 Jan 2024

   The term neurovascular coupling was coined in 2001 to indicate the interaction between brain activity and brain blood supply. While approximately a century ago it was believed that the vessels supplying blood to the brain and the brain cells function independently, neurovascular coupling evolved over time to highlight how the brain engages in regulating its own blood supply. Multiple mechanisms have been suggested and investigated that underlie neurovascular coupling.

   Here the investigators suggest a new mechanism for mediation of neurovascular coupling. By leveraging novel techniques, they showed that: 1) there are gaps in the astrocytic end feet that surround arterioles. 2) In these gaps, there are presynaptic axonal boutons that lie in the vicinity of underlying smooth muscle cells. 3) The axonal boutons are glutamatergic. 4) The smooth muscle cells participating in the junction with the boutons have glutamatergic receptors. 5) Activation of the glutamatergic receptors with small doses of glutamate, an indication of neural activity, is accompanied by calcium influx, which results in efflux of potassium, hyperpolarization and relaxation of smooth muscle cells, and vasodilation.

   The investigators also showed that profound release of glutamate, in pathologic conditions such as stroke, results in over-influx of calcium and contraction of smooth muscle cells, which dampens blood flow to already hypoxic brain cells and accentuates neural loss.

   The findings are very intriguing as they extend possible mechanisms underlying neurovascular coupling. The findings are presented by fascinating images of the cerebral vessels illustrating the gaps in astrocytic end feet and localization of neural-arteriolar smooth muscle junctions and related receptors.

   Several experimental models and tests were done to support study findings and conclusions. However, the main study findings are derived from experimental animal studies and cell cultures, which limits generalizability to human. Future human studies are needed to confirm expression of glutamate receptors in smooth muscle cells of cerebral small vessels. Moreover, more comparative studies are needed to elucidate the effect size of this mechanism in neurovascular coupling compared with other mechanisms including endothelial-derived vasodilators in both physiological and pathological conditions.

   View all comments by Shahram Oveisgharan

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References

News Citations

1. Aβ Acts Through Pericytes to Throttle Brain Blood Flow 21 Jun 2019
2. With Tau in Synapses, NO Neurovascular Coupling 21 Aug 2020

Paper Citations

1. Krimer LS, Muly EC 3rd, Williams GV, Goldman-Rakic PS. Dopaminergic regulation of cerebral cortical microcirculation. Nat Neurosci. 1998 Aug;1(4):286-9. PubMed.
2. Sándor P. Nervous control of the cerebrovascular system: doubts and facts. Neurochem Int. 1999 Sep;35(3):237-59. PubMed.
3. Iadecola C. The Neurovascular Unit Coming of Age: A Journey through Neurovascular Coupling in Health and Disease. Neuron. 2017 Sep 27;96(1):17-42. PubMed.
4. Joseph BK, Thakali KM, Pathan AR, Kang E, Rusch NJ, Rhee SW. Postsynaptic density-95 scaffolding of Shaker-type K⁺ channels in smooth muscle cells regulates the diameter of cerebral arteries. J Physiol. 2011 Nov 1;589(Pt 21):5143-52. Epub 2011 Sep 12 PubMed.
5. Moore CL, Nelson PL, Parelkar NK, Rusch NJ, Rhee SW. Protein kinase A-phosphorylated KV1 channels in PSD95 signaling complex contribute to the resting membrane potential and diameter of cerebral arteries. Circ Res. 2014 Apr 11;114(8):1258-67. Epub 2014 Feb 28 PubMed.

External Citations

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Further Reading

News

1. LOAD of Data Place Vascular Malfunction as Earliest Event in Alzheimer’s 8 Jul 2016