Online Inquiry

For Research Use Only. Not For Clinical Use.

Contact Us
  • Email:

Cholinergic Neurons Differentiation Service

Overview Our Services Published Data Applications FAQs Scientific Resources Related Services

Introduction of Cholinergic Neurons

Classification, Function, and Location of Cholinergic Neurons

The brain cholinergic system forms an extensive network of projection neurons, which innervate several brain areas. These neurons, defined as cholinergic because they contain the machinery for synthesizing and releasing acetylcholine (ACh) for neurotransmission, are grouped in nuclei. They include:

  1. The forebrain cholinergic neurons, forming a series of nuclei in the medial septum, the diagonal band of Broca, and the basal magnocellular nucleus of Meynert (NBM). They are among the largest and most complex neurons, with an axon length up to 50 cm and more than 1000 branch points. They are usually identified as Ch1, located in the medial septum, Ch2 in the ascending, Ch3 in the horizontal limbs of the diagonal band of Broca, respectively, and Ch4 in the NBM. The cholinergic neurons represent between 50 to 75% of the cells present in these nuclei and their projections form the main cholinergic input to the cerebral cortex, hippocampus, olfactory bulb, and amygdala. The widespread extension of the forebrain cholinergic system has been confirmed by analyzing, by magnetic resonance imaging, the parallel atrophy of the cholinergic neurons and the projecting cortical regions in patients affected by mild cognitive impairment (MCI).
  2. The large, aspiny cholinergic interneurons of the caudate nucleus and putamen.
  3. The cholinergic nuclei of the brainstem include the peduncle-pontine nucleus (PPN) Ch5, the dorsolateral tegmental nucleus (DLN) Ch6, the medial habenular nucleus Ch7, and the parabigeminal nucleus Ch8. Ch5 and Ch6 neurons project to the thalamus, hypothalamus, globus pallidus, forebrain cholinergic nuclei, and striatum, Ch7 neurons project to the interpeduncular nucleus, and Ch8 to the superior colliculus.

Schematic representation of some cholinergic pathways in the postnatal rat brain. Fig.1 Schematic representation of some cholinergic pathways in the postnatal rat brain.1

Cholinergic Neurons and Diseases

Post-mortem studies and in vivo neuroimaging investigations demonstrated that in Alzheimer's disease (AD), a loss of forebrain cholinergic nuclei from Ch1 to Ch4 is associated with cognitive deficits, ranging from MCI to dementia. A similar neuropathological situation characterizes alcoholic and post-traumatic dementia. Conversely, in Parkinson's disease (PD) without dementia, the forebrain cholinergic neurons are spared but there is a substantial loss of midpontine cholinergic neurons (C5 and C6), which contributes to the motor and sleep disturbances of this disease. Between these two conditions, there are Lewy body dementia (LBD), PD with dementia, and the Parkinsonian syndromes in which both forebrain and midpontine cholinergic nuclei are affected with the possible involvement of striatal cholinergic neurons.

Services at Creative Biolabs

Cholinergic Neurons Differentiation Service

As an industry-leading CRO company, Creative Biolabs has established a great reputation in neural differentiation models services. We have accumulated extensive professional experience during years of exploration. In addition, our Ph.D.-level experts team is also one of the strong guarantees of our high-quality services. If you are focusing on custom cholinergic neurons differentiation model, or you have any problems with our services, please don't hesitate to contact us for more information.

Our process begins with the cultivation of iPSCs using state-of-the-art protocols and growth factors. These pluripotent cells are reprogrammed to differentiate into cholinergic neurons under strictly controlled in vitro conditions which closely mimic the internal environment of the human body. This complex differentiation process typically lasts several weeks.

After the differentiation process, we perform a series of quality controls. We assess the efficiency of the differentiation through gene expression with RNA sequencing and via neuron immunostaining for specific markers of cholinergic neurons. These methods ensure the accuracy and quality of the generated neurons.

Therefore, our cholinergic neuron differentiation service ensures you receive ready-to-use, high-quality cholinergic neurons for your cell-based research. We work closely with clients to discuss their specific requirements and customize our service to meet their precise research needs. We also offer flexibility in our services, including but not limited to:

Services Descriptions
Alzheimer's Disease Models Service Creative Biolabs is an experienced custom in vitro CNS disease modeling services provider. Our platform is now mature in offering various in vitro CNS disease services, including Alzheimer's disease models.
High Content Screening Service In neuroscience, HCS has been used in two main areas: neuroregeneration and neuroprotection. Our HCS service, which primarily uses a range of primary neurons, contributes to the development of anti-neurological drugs through the quantitative analysis of complex cellular phenotypes.
STEMOD™ Advanced Drug Discovery Service We have developed a comprehensive technology platform to provide one-stop CNS drug discovery services. Our platform has advanced neuroscience ex vivo models, neuroscience assay techniques, and neuroscience research tools.

Published Data

P. A. Goldsteen et al. used human pluripotent stem cell (hPSC) technology to develop an in vitro model of human peripheral cholinergic neurons. They established a robust cholinergic neuron differentiation scheme for the study of respiratory neuroeffector communication. The scheme used dual SMAD inhibition and Wnt activation to generate p75+ -HNK1+ NCC precursors. Subsequently, BDNF was used to priming vagal NCC to mature and functional peripheral cholinergic neurons.

As shown, the neuronal network was clearly visible at day 25 of differentiation and further expanded and became denser over time. Expression of VAChT and SLC18A3 confirmed the cholinergic phenotype of the neurons.

Immunofluorescence images of differentiating cholinergic neurons over time and gene expression of cholinergic neuronal development over time.(Goldsteen, P. A., et al., 2022)Fig. 2 Immunofluorescence images of differentiating cholinergic neurons over time and gene expression of cholinergic neuronal development over time.2


Our advanced, state-of-the-art cholinergic neuron differentiation service is primarily applied in two major areas:

  • Neurodegenerative Disease Research: Our service aids in creating powerful models for studying neurodegenerative disorders like Alzheimer's and Parkinson's disease. These conditions are associated with a deterioration in cholinergic neurons, and hence using these differentiated neurons allows for a comprehensive understanding of the disease's mechanisms.
  • Drug Discovery and Testing: We offer our services to pharmaceutical companies for effective drug discovery and toxicology testing. Testing potential medications on cholinergic neurons first ensures that the drugs are safe and efficacious before they are subjected to clinical trials.

Leveraging a well-defined protocol, we provide robust cholinergic neuron cells with a high differentiation efficiency, which are ideal for both high-content screening and high throughput screening. The offering is fully customizable to meet our clients' unique requirements. We ensure strict confidentiality and adherence to regulatory standards in all our services.


  • Q: Do you guarantee the purity of the differentiated neurons?
    A: Yes, we do. Our experienced and skilled professionals employ stringent measures throughout the differentiation process to ensure a high degree of purity and minimize the risk of contamination. However, the exact level of purity can vary depending on different factors, including the characteristics of the initial pluripotent stem cells.
  • Q: How do you stay abreast of the latest advancements and innovations in cholinergic neuron differentiation techniques and incorporate them into your service offerings?
    A: We are dedicated to staying at the forefront of cholinergic neuron differentiation techniques and innovations by actively monitoring the latest research developments, attending scientific conferences, and collaborating with leading experts in the field. We continuously evaluate emerging technologies and methodologies to enhance our service offerings and incorporate cutting-edge advancements into our protocols.
  • Q: Is the service available worldwide?
    A: Yes, we provide our services globally. However, the delivery timelines and shipping conditions may vary depending on the requirements of specific regions or countries.
  • Q: How to ensure reproducibility across experiments?
    A: Reproducibility is central to our approach. We maintain strict adherence to standardized protocols and use validated reagents from reputable sources. Additionally, our team undergoes regular training to ensure consistency in technique and execution, minimizing variability and enhancing the reliability of results across experiments.

Scientific Resources


  1. Shabani, Z., et al. An Overview of Nicotinic Cholinergic System Signaling in Neurogenesis. Arch Med Res. 2020, 51(4): 287-296.
  2. Goldsteen, P. A., et al. "Differentiation and on axon-guidance chip culture of human pluripotent stem cell-derived peripheral cholinergic neurons for airway neurobiology studies." Frontiers in Pharmacology 13 (2022): 991072.

For Research Use Only. Not For Clinical Use.