Network vs Individual Bursting Neurons

Motor Neurons and MEA
Dysregulated bursting is at the root of many motor neuron/neuromuscular junction disease. ArunA Biomedical teaming with Axion Biosystems have generated relevant bursting data from our Mouse Motor Neurons cultured on Axion-Bioystem’s Maestro MEA.

Figure: Mouse Motor Neuron Network Modulation by Bicuculline-ckeck out the entire presentation to learn more: GFP+ Motor Neurons: Development and in-vitro Functional Assessment on Microelectrode Arrays

Protocol User’s Guide for Culturing Motor Neuron on MEA(pdf – 679Kb)

Name Catalog # Type Species Applications Size Price
Motor Neurons-GFP+ Quick Start Kit mMN7205.QS Primary Neurons M Cell Assays 750,000 $349
Motor Neurons-GFP+ HTS Kit mMN7205-HTS Primary Neurons M Cell Assays 4 X 750,000 $989
GDNF (Human, Mouse) PR27022-2 Protein H; M 2 ug 10 ug $108 $205
AB2™ Basal Neural Medium AB27011.3 Cell Growth Media H; M Cell Assays 500 ml $69

We will continue providing you content we believe important. Should you have questions, do not hesitate to contact us. Thank you and we stand ready to serve you and your team.

Pete Shuster-CEO and Owner, Neuromics, 612-801-1007,

eSC Derived hNP1 Neural Progenitors Astrocytic Differentiation

Protocol for Driving hNP1TM Human Neural Progenitors to Astrocytes

There is a great demand for an easy way to generate human astrocytes in culture. I am pleased to present a protocol for differentiating our hNP1 Cells to Astrocytes. This comes from my friend Dr. Steve Stice and his team at ArunA Biomedical and University of Georgia: Majumder A, Dhara SK, Swetenburg R, Mithani M, Cao K, Medrzycki M, Fan Y, Stice SL. Inhibition of DNA methyltransferases and histone deacetylases induces astrocytic differentiation of neural progenitors. Stem Cell Res. 2013 Jul;11(1):574-86. doi: 10.1016/j.scr.2013.03.003. Epub 2013 Apr 2.

These enriched non-transformed human astrocyte progenitors will provide a critical cell source to further our understanding of how astrocytes play a pivotal role in neural function and development. Human neural progenitors derived from pluripotent embryonic stem cells and propagated in adherent serum-free cultures provide a fate restricted renewable source for quick production of neural cells; however, such cells are highly refractive to astrocytogenesis and show a strong neurogenic bias, similar to neural progenitors from the early embryonic central nervous system (CNS). We found that several astrocytic genes are hypermethylated in such progenitors potentially preventing generation of astrocytes and leading to the proneuronal fate of these progenitors. However, epigenetic modification by Azacytidine (Aza-C) and Trichostatin A (TSA), with concomitant signaling from BMP2 and LIF in neural progenitor cultures shifts this bias, leading to expression of astrocytic markers as early as 5days of differentiation, with near complete suppression of neuronal differentiation.

Images: Morphology and gene expression after 15 and 30 days of differentiation of cells with astrocytic treatment. Bright field images of hNP cells differentiated (A) with or (B) without astrocytic treatment. A and B compare morphology of cultured cells in treated vs. untreated differentiation at 15 days. Treated and untreated cells were cryopreserved at d6 and subsequently thawed and cultured for an additional 9 days. Flow cytometry analysis to determine percent of GFAP+ and S100B+ cells at d15 of differentiation. Data is presented as histograms for (C) GFAP and (D) S100B with corresponding immunoreactive cells in insets from a parallel culture. Immunocytochemistry detects expression of (E) GFAP with S100B (inset showing distinct staining for both markers), (F) GFAP with GLAST, and (G) GFAP with ALDH1L1 at d30 of differentiation.

The Protocol:  For astrocytic differentiation of hNP cells, neuronal differentiation media were supplemented with BMP2 (20 ng/mL) and combinations of Aza-C and TSA; Aza-C (500 nM), TSA (100 nM) and BMP2 (20 ng/mL) for 2 days, with one complete media change in between, followed by differentiation media supplemented with BMP2 but not with Aza-C or TSA. Cells were harvested prior to analysis at 5, 15 or 30 days of treatment or for cryopreservation to d6 or d10 of differentiation. For cryopreservation, cells were
dissociated with Accutase™ and frozen in differentiation media containing 10% DMSO. Viability was assessed at 30 days in Aza-C and TSA treated cultures by trypan blue exclusion, and datawas acquired using a Cellometer Auto T4® (Nexcelom Biosciences).

I will keep you updated on new differentiation protocols for our potent, pure and widely used hNP1 Human Neural Progenitors to new phenotypes.

Musculoskeletal Disorders-Stem Cell Based Drug Discovery

A common Neuromics’ theme is harnessing the power of cellsTM. The raw cost of the cells are often the biggest consideration. We encourage our customers to focus on true costs. These include the # of cells (how many times can they be passaged?), culture viability (how long do the cells live) and bioactivity (how closely do cultures mimic in vivo behavior?). I would like to present a presentation and publication confirming our competitive advantage when analyzing true costs.

Setting a higher bar for Neuron-Glial Based Assays!

Dr. Randen Patterson and his team at UC Davis have developed new culturing techniques using our e18 Rat Primary Hippocampal Neurons. They have developed a protocol that allows for culturing of E18 hippocampal neurons at high densities for more than 120 days. These cultured hippocampal neurons are (i) well differentiated with high numbers of synapses, (ii) anchored securely to their substrate, (iii) have high levels of functional connectivity, and (iv) form dense multi-layered cellular networks. We propose that our culture methodology is likely to be effective for multiple neuronal subtypes–particularly those that can be grown in Neurobasal/B27 media. This methodology presents new avenues for long-term functional studies in neurons. This is good news indeed: Todd GK, Boosalis CA, Burzycki AA, Steinman MQ, Hester LD, et al. (2013) Towards Neuronal Organoids: A Method for Long-Term Culturing of High-Density Hippocampal Neurons. PLoS ONE 8(4): e58996. doi:10.1371/journal.pone.0058996.

We will continue to raise the bar. Better cultures=lower costs and better outcomes!

Featuring Dr. Richard Rogers

Obesity Energy, Thermogenesis and Appetite

Dr. Richard Rogers

Dr. Richard Rogers

Obesity and its evil twin, diabetes, are rapidly becoming our #1 health epidemic. Today 10% of all medical costs in the U.S. are dueto an overweight population, and this percentage is growing rapidly. Today, the breakdown is about $1500 per year in medical costs for obese versus normal weight individuals. This translates into more than $145 billion spent annually.
Given the size of the epidemic, a growing focus for my company is providing reagents to researchers who study bioprocesses involved in energy metabolism. This includes researchers studying what happens to energy expenditure when the “fuel tank” is full and also empty. Both states could give clues as to why we overeat.

In my routine follow up with researchers using our reagents, I started to get an appreciation on how these complex energy pathways are being unraveled and better understood. That appreciation forms the roots of this “News Behind the Neuroscience News” story. It is a story that has the hormones Leptin and TRH playing a starring role supported by hindbrain neuro/glial-circuitry and brown adipose tissue (BAT).

The Rogers Lab

I became aware of the Richard Rogers Lab in my follow up with Montina Van Meter checking on our  LepRb/OBRb antibody. She shared that it was giving them results better than most others they has used. She then gave me an overview of her research involved with  the control of feeding behavior and energy utilization including thermogenesis (“heat generation”) catalyzed in BAT.  Cool…this was a lab we wanted to make sure we served and served well.

Tina not only kept me informed on how our reagents were working, but also generously alerted to me publications referencing use of our reagents:
·        Maria J. Barnes, Richard C. Rogers, Montina J. Van Meter and Gerlinda E. Hermann. Co-localization of TRHR1 and LepRb
receptors on neurons in the hindbrain of the rat.
 doi:10.1016/j.brainres.2010.07.094…Included are excellent images of stained LepRb (OB-Rb)-(Dilution 1:500) and  GAD1-Dilution (2ug/ml) expressing neurons localized in loose clusters of cells in the DMN, NST, and the VLM…
·        Hung Hsuchou, Yi He, Abba J. Kastin, Hong Tu, Emily N. Markadakis, Richard C. Rogers, Paul B. Fossier, and Weihong Pan. Obesity induces functional astrocytic leptin receptors in hypothalamus. Brain, Mar 2009; doi:10.1093/brain/awp029…unique sequence of ObRb at its cytoplasmic tail (CH14104, Neuromics, Edina, MN, USA). This antibody was raised
against rat ObRb…

I found this research to be unique and intriguing. This led me to an interview with Dr. Rogers. Here is his backstory.


Dr. Rogers credits serendipity as a driving force in his interest in Neuroscience. It started with a bike ride and chance introduction with a ham radio operator when he was a youngster. This catalyzed his interest in electronics and circuitry.

This interest morphed to a passion for Neuroscience (circuits and signaling). He entered the first college program devoted to Neuroscience studies at UCLA.  He received his Ph. D. in 1979. His post-doc focused on digestive regulation. Here, he  investigated the neural circuitry involved in the normal control of gastric function.


In collaboration with his wife Dr. Gerlinda Hermann, his work evolved to solving the mystery of why we don’t eat (abnormal gastric function). What causes gastro-intestinal shutdown?  The breakthrough was their ability to show cross-talk between the immune system and nervous system. This research is a foundation for the discovery of therapies for sufferers of appetite shut down cause by cancer therapies and certain immune related pathologies.

The main culprit is TNF-α. The blood level of this peptide is elevated as a consequence of immune activation caused by infection, cancer, radiation exposure and chronic autoimmune disease. The breakthrough was showing that  TNF-α receptors are on neurons in the brainstem that control gastric functions, including emesis.  Neurons in the nucleus of the solitary tract (NST) respond to TNF-α greatly increasing the sensitivity of gastric vagal control circuitry.  This causes emptying of the gut to dramatically slow,l leading to nausea, vomiting and cessation of appetite.

Currently, they are delving into the complexities of  TNF-α signaling processes. This includes the role of astrocytes and glia.

See: Gerlinda E Hermann and Richard C Rogers.  TNF activates astrocytes and catecholaminergic neurons in the solitary nucleus: implications for autonomic control. doi: 10.1016/j.brainres.2009.03.059.

 Energy,Obesity and Thermogenesis-Active Astrocytes

Recently, the lab took another road less traveled. Dr. Gerlinda Hermann discovered interesting aspects of leptin and thyrotropin releasing hormone (TRH) signaling.  This research looks at signaling in thermogenesis and feeding behavior. A most interesting aspect includes conclusions concerning the role of astrocytes.  Their colleague Dr. Weihong Pan  showed that adult obese mice, (2 months after being placed on a high-fat diet) showed a striking increase of leptin receptor (+) astrocytes, most prominent in the dorsomedial hypothalamus and arcuate nucleus. Agouti viable yellow mice with their adult-onset obesity showed similar changes, but the increase of leptin receptor (+) astrocytes was barely seen in ob/ob or db/db mice with their early-onset obesity and defective leptin systems. The results indicate that metabolic changes in obese mice can rapidly alter leptin receptor expression and astrocytic activity, and that leptin receptor is responsible for leptin-induced calcium signalling in astrocytes. This novel and clinically relevant finding opens new avenues in astrocyte biology (doi: 10.1093/brain/awp029).

Non-shivering thermogenesis usually occurs in BAT. It uncouples the ATP energy producing process by generating heat rather than driving the conversion of ADP to ATP. This creates an ingenious way to untangle complex processes related to Leptin signaling. What happens when there is sufficient energy for the thermogenic process? Conversely, what happens when there is insufficient energy?

 Leptin and TRH act synergistically in the hindbrain to drive thermogenesis. However, in a starving condition there is a subsequent drop in Leptin and thermogenesis. Behind these simple facts are complex processes that occur in the hindbrain. The team is providing important insights including location of events and relevant signaling molecules. (doi:10.1016/j.brainres.2010.07.094).

The Future-Caged Compounds and Live Cell Signaling

Caged compounds are bioactive molecules attached to photolabile groups, that release the active component on contact with photons of the right energy level – the process of photolysis. Photolysis is now widely applied in biology to induce neurotransmitter and otherm ligand-receptor interactions in conditions that are otherwise subject to poor diffusional access and receptor desensitization, as well as for labile ligands.

This novel technology affords Dr. Rogers and his team the capability do live cell imaging of calcium signaling. From these they will help us gain a deeper understanding of what is happing and where. Specifically, we will more exactly learn the role that astrocytes and glia play in controlling the role of   Leptin and related signaling molecules in controlling energy, metabolism and feeding behavior. This could lead to important target for future therapies.

Umbilical-Cord Matrix Stem Cells and Cerebral Ischemia

I am winding down on the stem cell story for now as later in the month I will be featuring my good friends at University of Sherbrooke and their research in the area of chronic pain.

I did want to highlight yet another potential application for stem cells. For this, we send kudos to Dr. Yan Xu and his colleagues at University of Pittburgh for their findings on inflammatory response in Golbal Ischemia. Their work was recently published:

Aaron Hirko, Renee Dallasen, Sachiko Jomura, Yan Xu. Modulation of Inflammatory Responses after Global Ischemia by Transplanted Umbilical-Cord Matrix Stem Cells. Stem Cells First published online August 21, 2008; doi:doi:10.1634/stemcells.2008-0075

Secondary to Cardiac Arrest is Brain Damage do to lack of blood flow. This is marked by a delayed loss of Neurons in CA1 hippocampus region of the brain due to inflammatory response.

The story timeline of this response is good then bad with interesting twists. The delay in neuronal loss is linked to initial inflammation. It involves both reactive astrocytes (astrocytosis) and glia. Delaying the loss is, of course, good.

…But then, the reactive astrocytosis and related glial scarring cause a physical and biochemical barrier to regeneration of neurons…a bad thing. Protecting the microglia is a good thing, because they these cells serve as scavengers for clearing the cellular debris. They can also secrete a variety of cytotoxic and protective chemicals.

The wow factor in this research is that  implanted rat umbilical-cord matrix (RUCM) cells can provide partial protection against neuronal injury in rat brains. Rats treated with RUCM cells three days prior to an 8-min CA had only 25-32% neuronal loss in the hippocampal CA1 region compared to the typical 50-68% neuronal loss observed in the untreated or the vehicle-treated animals. This could be due to to the favaorable modulation of the “good-bad” inflammatory response.

The good news in the search for therapies for stroke and cardiac arrest victims is combined, stem-cell-like RUCM cells offer protection against neuronal injury after global cerebral ischemia by enhancing the survivability of the astroglia in the selectively vulnerable regions.

We are pleased that the research team used our GFAP antibody as an marker for astrotytic in their studies.

Spinal Cord Injury and Ependymal Cells

This research sheds light on the natural regeneration that occurs in the area of damaged spinal cord tissue. The surprise here is the role that Edendymal Progenitor Cells are playing in repair mechanism vs Neural Stem Progenitors. 

In this publication, researchers have employed genetic fate mapping to characterize a candidate neural stem cell population in the adult spinal cord and show that close to all in vitro neural stem cell potential resides within the population of ependymal cells. Ependymal cells give rise to a substantial proportion of scar-forming astrocytes as well as to some myelinating oligodendrocytes after spinal cord injury. Modulating the fate of ependymal cell progeny after injury could potentially promote the generation of cell types that may facilitate recovery after spinal cord injury.

Meletis K, Barnabé-Heider F, Carlén M, Evergren E, Tomilin N, et al. (2008) Spinal Cord Injury Reveals Multilineage Differentiation of Ependymal Cells . PLoS Biol 6(7): e182 doi:10.1371/journal.pbio.0060182