A Guide to Neuronal Cell Markers

Neurons are the cells that give rise to our perceptions, memories, and behaviors. They do so by communicating through dense, intricate networks of synapses, the neuron-to-neuron junctions through which neurons transmit chemical (neurotransmitter) and electrical signals.

Nearly 100 billion neurons populate the human brain during development. During adulthood, this number is maintained through cell death and replenishment via neurogenesis. Given the vast diversity of this massive population, this guide focuses only on general neuronal markers, or those that can be detected irrespective of anatomical location in the brain, including markers of mature neurons, neuronal activity, synapses, and neurodegeneration. Not reviewed are markers of neuronal subtypes (e.g. excitatory neurons and interneurons, neurotransmitter markers, etc.). 

Mature Neuronal Markers

Many markers can be used to label mature neurons throughout the brain. Not only do these markers label neurons, they label specific parts of neurons such as the cell body, nucleus, and processes (axons and dendrites). Neuron-specific enolase (NSE, encoded by ENO2), for example, is a neuron-specific marker that labels neuronal cell bodies. Neuronal nuclei can be visualized with NeuN (a transcription factor encoded by RBFOX3). Processes have many more markers. TUJ1, neurofilament (NEFL), and tau (MAPT) mark axons, whereas MAP2 labels dendrites. These popular markers are suitable for immunohistochemical labeling of neurons in all regions of the brain.

Image: This figure highlights the general development of neurons and notable protein markers.

Synaptic Markers

When you zoom in on dendrites, stubby protrusions can be seen lining the entire length of the processes. These structures, known as dendritic spines, receive chemical (and sometimes electrical) signals from the axonal terminal of other neurons. These junctions are the basis of neuronal communication. As such, they have been studied deeply, leading to the identification of many synaptic markers.

Location in the presynaptic side of the synapse, syntaxin (STX1A), SNAP25, synaptotagmin (SYT), SV2A, and synaptophysin (SYP) all regulate the release of neurotransmitters from axonal terminals into the synaptic cleft. Members of the DLG and SHANK gene families (DLG1, DLG2, DLG3, DLG4, SHANK1, SHANK2, SHANK3), by contrast, serve as scaffolding proteins in dendritic spines, helping to organize receptors and complexes on the postsynaptic membrane. The neuroligin (NLGN) gene family (NLGN1, NLGN2, NLGN3, NLGN4, NLGN5) bridges the gap between presynaptic and postsynaptic membranes by ligating with β-neurexins. Other synaptic markers include neuronal pentraxin 2 (NPTX2), a synaptogenic protein expressed rapidly in response to neuronal depolarization (activation of the postsynaptic neuron); protein phosphatase 1 regulatory subunit 9B (PPP1R9B), a regulatory subunit of protein phosphatase-1 catalytic subunit; and neurogranin (NRGN), an important regulator of calcium signaling in dendritic spines.

Markers of Neuronal Activity

Classical behavioral studies using protein synthesis inhibitors demonstrated that protein synthesis is a requirement of learning, synaptic plasticity, and memory formation. These studies led to the hypothesis that activity-dependent immediate-early gene (IEG) expression plays a critical role in these processes. They also spurred a search for novel IEGs after the initial discovery of c-Fos (FOS). Such markers could be used to immunohistochemically label “active” neurons in preserved brain samples.

Large gains in the understanding of IEGs were made in the 90s when over a dozen were discovered using various screening methods. Among these newly discovered IEGs were FOSB, JUN, JUNB, and NR4A1 as well as EGR1, EGR2, and EGR3 of the EGR gene family, all of which encode transcription factors; ARC and HOMER1A, which encode postsynaptic proteins; RHEB, RSG2, SNK, and COX2, which encode intracellular signaling proteins; BDNF, NHBA, PLAT, and NP2, which encode secreted factors; and the membrane protein neuritin (encoded by NRN1). Despite the discovery of all of these IEGs, c-Fos is the most commonly used marker of neuronal activity.

Neurogenesis Markers

New neurons can be generated during adulthood in a process called neurogenesis. This process takes place within the dentate gyrus of the hippocampus, a structure that is associated with learning and memory; indeed, neurogenesis is thought to contribute to these processes throughout adulthood.

Neurogenesis begins with a GFAP⁺Nestin⁺BLBP⁺SOX2⁺ stem cell progenitor, called type 1 cells. These cells develop into type 2 cells, a rapidly proliferative cell type that downregulates SOX2 and upregulates EOMES (TBR2), PROX1, NEUROD, NCAM1, and DCX. These cells downregulate Nestin as the transition to type 3 cells, the last stage in the precursor stage of neurogenesis.

When these progenitors exit the cell cycle, they begin expressing mature neuronal markers, including NeuN (RBFOX3) and calretinin (CALB2). As these cells mature further and form synapses, they switch from expressing calretinin (CALB2) to calbindin (CALB1), both of which act to buffer calcium. Within a few weeks, the newly formed cells are indistinguishable from their mature, granule-cell neighbors.

Neurodegeneration Markers

Just as new neurons are formed throughout adulthood, older neurons can degenerate and eventually die. Such processes underlie a variety of conditions, including Alzheimer’s disease, Parkinson’s disease, and even mild neurocognitive impairment.

One way to investigate neurodegeneration is to look at the loss of axonal, dendritic, and synaptic markers. The proteins encoded by MAP2, NEFL, TUJ1, SYP, and DLG4 (more commonly known as postsynaptic density 95, or PSD95) are frequently used to quantify, by immunohistochemistry, the loss of neuronal processes across neurodegenerative conditions. Virtually all of the synaptic markers discussed above, however, can serve as neurodegeneration markers, and many have been observed at elevated concentrations in the cerebrospinal fluid of patients of various neurodegenerative conditions.

Other markers are more specific to neurodegenerative conditions. For example, the protein products of APP and MAPT — amyloid-β and tau — are important pathological hallmarks of Alzheimer’s disease. The aggregation of these proteins indicates progression to Alzheimer’s disease. Presinlin 1 and 2 (PSEN1 & PSEN2) as well as beta-secretase 1 (BACE1), which regulate the generation of amyloid-β, are mutated in early-onset (familial) Alzheimer’s disease.

Table of Neuronal Markers

The table below lists characteristic neuronal proteins as described by review literature. The list includes a variety of marker types, including transcription factors, membrane proteins, secreted factors, signaling proteins, and structural proteins. Accompanying each marker are links to relevant antibodies and ELISA kits that can be used to detect neurons in vitro and in vivo. The associated products are offered by a variety of manufacturers and can serve as a useful reference for neuronal characterization.

Note: *Some markers are protein isoforms, multi-subunit protein complexes, or protein families composed of several distinct genes. Information on Protein Type, Localization, and Size (kDa) obtained from UniProt.org (for human genes only). 

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