Hello Swati.

This question is a complex and asking varying aspects of 1) ongoing works of revealing evidences of anxiolytic drug effects on the nervous system, 2) wrong term for anxiety/stress biomarkers (they are physiological or psychological or clinical), and 3) complexities of the medicinal plant extract you mentioned.

I assume that the anxiety/stress markers mentioned are pointing out the genes expressed upon antidepressant treatment in various sources of nervous cells/tissues in vitro or in vivo in many reports. These works are not finding antidepressant effects, but revealing additional effects occurring as side effects on the nervous system. The effects vary from positively neurogenic and neuronal maturation or negatively breaking synapses or transformation of glia to neuronal phenotype differentiation. Thus, it has to be resolved with many more works with different sources of neurons or glia in vitro and in vivo.

About plant extract, it has so many compounds, you would expect cell proliferation (neurogenic), cell death (breaking sysnapses), or accumulation or abolishment of ROS within the cells depending on the concentration used. And no antidepressant markers were found so far as I discussed in vitro. It would be impossible to show pure anxiolytic effects with the cell lines or primary cells in vitro.

About cell lines, they are derived from different sources, you would obtain varying results depending on the cell lines or primary cells used.

Think more about the experiments. If you use biomarkers of the genes expressed as in conflicting reports, you would look at thepositive ornegative neurogenic effects. Then, your plant extract is functionally not anxiolytic, but neurogenic or neuro-dedegrading agent.

Best wishes.

Dear Sir,

Thank you for your prompt response and clarification on the topic. I've done some priliminary work on the mouse models of anxiety including behavioral analysis and neurotransmitter quantitation in different brain regions. Can you suggest some in vitro experiments on cell lines or in mice to support this work?

Hi Swati.

As I discussed, no obvious anxiolytic markers are found yet. The claimed biomarkers reported are either toxic effects related to inflammatory or stress-related genes or the unexpected side effects of the drugs, showing enhancement of neuronal maturation or opposite effects to that. I do not know whether you can find such anxiolytic markers using cells in vitro. The physiological responses regulated at the level of complex body never happen in a single cell population in vitro. Because the states of anxiety or release of anxiety are consciousness dimension, not the cell materials.

However, if you insist, you could use PC12 cells for PNS model (adrenal gland-derived neuroendocrine properties) and N2a cells for CNS (neural stem cell properties, requiring cell differentiation induction), but IMR32 cells are mixed origins of cell population (neuron and fibroblast properties).

Regards.

What kind of markers should I look after in these cell lines? Also I was suggested to check for c-fos expression by one of the neuroscientist from TIFR, Mumbai.

Hello Swati.

The question you asked is only one of many worries about your works predicted.

Increased translocation of c-fos into neuronal cell nuclei can occur when you give cells under stress with your plant-derived mixed compounds, but not by the anxiolytic agent(s) in the extract. It would be more obvious if you test another plant extract that has no anxiolytic agent(s) and known to be non-anxiolytic along with your anxiolytic extract in your experiment.

Sorry, I already said there is no clear anxiolytic biomarkers established by any research due to the just above-mentioned reason, or due to the toxic effect of tested drugs.

Induced c-fos expression is found either in activated neurons by NGF or in the condensing chromatin of apoptotic neurons, but not by the anxiolytic action. So, how do you know whether your extract has toxic effects on neurons by increasing ROS, leading to cell death, and has NGF-like stimulant in the extract, activating neurons regardless of anxiolytic effects.

Please read the first reply more carefully to understand what I described, and read the meaning, but not the sentences.

Regards.

Thank you so much sir for you valuable suggestion and information. Looking forward to more communication and knowledge exchange in future.