Case study

Targeting TRP channels for the treatmentof eye diseases

Tools:
Port-a-Patch

Dr. Stefan Mergler
featured by Nanion Technologies
Stefan Mergler is a researcher and lecturer

are associated with night blindness. Bestrophin-1 channels,

as well as a group leader at the Charité

implicated in Best vitelliform macular dystrophy, play a

University Hospital in Berlin, Germany. His

significant role in the retinal pigment epithelium. The CFTR

research focuses on the role of TRP channels

channel, involved in transepithelial transport, along with ClC-

in eye diseases, aiming to develop effective

2 channels, critical for chloride transport, have been linked totherapies for conditions such as dry eye

retinal degenerations. In the anterior segment of the eye, adisease and retinoblastoma.

number of temperature-sensitive TRP channels such as TRPV1and TRPM8 are specifically involved in dry eye disease, whichis the most common eye disease in ophthalmology. Notably,
TRPV1 inhibitors have also been shown to offer avenues fordeveloping novel treatments for such an ocular condition.

Eye diseases, ranging from common conditions like cataracts
and glaucoma to complex disorders such as maculardegeneration, affect millions worldwide, significantly
impacting vision and quality of life. These conditions vary insymptoms and severity, with some leading to gradual vision
loss and others causing acute blindness.
Ion channels play critical roles in various aspects of eyehealth and disease. Ion channels in ocular tissues, including
the cornea, conjunctiva, lens, retina, and retinal pigment
epithelium, are crucial for maintaining corneal transparency,
intraocular pressure, and nutrient supply. They also play apivotal role in the light transduction process in photoreceptors
as well as release of pro-inflammatory cytokines.
Dysfunctions in specific ion channels are linked to various eye
diseases and therefore, these channels represent potential
targets for pharmacological intervention. Cyclic NucleotideGated (CNG) channels have been linked to conditions
like retinal degeneration (e.g. retinitis pigmentosa) andachromatopsia in the posterior segment of the eye. L-type
Ca2+ channels CaV1.4, crucial for neurotransmitter release,

Nanion's Port-a-Patch platform
is a turn-key miniaturized patch-clamp systemenabling the user to rapidly generate high quality
data, regardless of experience. Behind its small
and compact appearance lies sophisticatedtechnology, producing high-quality measurementswith giga-seals and high success rates.

Among a few available methods, the Nobel Prize-awarded
patch-clamp technique is most frequently used to investigate
the biophysical properties of ion channels. Manual patchclamping has consistently been the preferred standard for ion
channel studies, as it provides a level of detailed data that
virtually no other techniques can match.

“The Port-a-Patch has significantly accelerated our research.
Since 2010, we’ve published more than 20 papers with it.
That speaks for itself.”
Dr. Stefan Mergler, Charité University Hospital

However, while it’s a powerful tool, the conventional patch-

Launched in 2003, the Port-a-Patch instrument has literallyclamp technique is often considered to be complicated, time-

revolutionized the field of electrophysiology. This small

intensive, laborious, and not conducive to high-throughput

benchtop semi-automated planar patch-clamp device has

experimentation. It necessitates considerable training andenabled scientists to conduct their ion channel researchtechnical skills and involves using a complex set of equipment

more easily and allows users to rapidly generate high-quality– factors that are regarded as drawbacks or limitations of thisdata, regardless of experience. Since its launch, the Port-a-

technique.

Patch has been widely adopted around the world – over 200
papers have been published using the Port-a-Patch to date.

It’s encouraging that over the past twenty years, a few

Numerous groups have successfully used the Port-a-Patch tocompanies have made strides in creating an automatedstudy both voltage- and ligand-gated ion channels in cell lines,

version of the patch-clamp technique (APC). APC has

primary cells, stem cell-derived cells, and even organelles.

addressed many of the challenges inherent in manual patchclamping. Current APC systems are capable of simultaneously

One of these groups, led by Stefan Mergler, has long beenmeasuring currents from a range of 1 to 384 cells. These

studying the role of TRP channels in eye diseases, aiming

advanced systems are user-friendly and often come withto develop effective therapies for conditions like dry eye

automated data analysis features, which considerably

disease, uveal melanoma or retinoblastoma. Since 2008, thereduces the time researchers need to spend on data analysis.

Port-a-Patch has been extensively employed in Mergler’s lab,

L-carnitine inhibits TRPV1 activation in human corneal epithelial cells. TRPV1 channel can beselectively activated by capsaicin, hypertonicity, or heat (>43 °C), and blocked by L-carnitine.
Three different methods were used to validate the inhibitory effect of L-carnitine, such as
single-cell fluorescence calcium imaging, cell volume measurements and planar patchclamp.1

aiding in the accurate recording of TRP channel activity andkey factors in dry eye is an increase in tear osmolarity, which

complementing techniques like calcium imaging. Dr. Mergler

is known to activate the osmosensitive TRPV1 channel.

notes that “the Port-a-Patch has significantly accelerated our

L-carnitine has previously been demonstrated to be an

research. Since 2010, we’ve published more than 20 papers

osmoprotectant against hyperosmotic stress in corneal

with it. That speaks for itself.”

epithelial cells. The topical use of L-carnitine has been shown
to result in rapid and consistent improvements in the symptoms

Over the years, Stefan Mergler’s lab used the Port-a-Patchof dry eye patients. By combining Port-a-Patch-based

to study thermo-TRPs (mainly TRPV1, TRPV4 and TRPM8electrophysiology with calcium imaging and cell volume

channels) in a variety of cell lines and primary cells, including

measurements, Dr. Mergler and colleagues have shown thathuman corneal endothelium cell line HCEC-12 and epithelial

L-carnitine suppresses capsaicin- and hypertonicity-induced

cell line HCE-T, pancreatic neuroendocrine tumor cell line

TRPV1 activation by blocking cell volume shrinkage. This effect

BON-1, human retinoblastoma cell line WERI-Rb1, human

may be of therapeutic value as TRPV1 activation is known toconjunctival epithelial cell lines HCjE and IOBA-NHC, humanlead to proinflammatory cytokine release and ocular surface

corneal keratocytes HCK, human uveal melanoma cell line

inflammation in dry eye. Furthermore, TRPV1 inhibition has

92.1, human pterygial cells hPtEC, and embryonic mouse

been reported to reduce ocular pain as well as tissue fibrosis in

hypothalamic cell line mHypoE-N41.1-20

dry eye disease.1

In their recent study, the group employed the Port-a-Patch

Previously, the group also showed that L-carnitine, acting

to examine the effect of L-carnitine on TRPV1 activation in aas an osmoprotectant, suppressed hypertonicity-induced

human corneal epithelial cell line (HCE-T) in the context of dry

TRPV1 activation in human conjunctival epithelial cells andeye syndrome, a widespread ocular disorder characterized

human corneal keratocytes (HCK).2 They demonstrated thatby a stinging or burning sensation in the eyes.1 One of the

L-carnitine inhibited TRPV1 currents in HCK cells, leading to

Dr. Stefan Mergler performs patch-clamp experiments using the Port-a-Patch at Charité University Hospital.
His research focuses on the role of TRP channels in eye diseases, aiming to develop effective therapies forconditions such as dry eye disease and retinoblastoma. Picture by Wiebke Peitz (Charité newspaper).

“The Port-a-Patch is a very capable system that has been
instrumental in our research. Additionally, it is much easier to use
than a manual patch-clamp rig, making it an excellent tool not
only for research purposes but also for teaching and learning
patch-clamp technique.”Dr. Stefan Mergler, Charité University Hospitalreduced keratocyte transdifferentiation into myofibroblasts.

References

These results suggest that blocking TRPV1 in keratocytes with1.

Lucius A, Chhatwal S, Valtink M, Reinach PS, Li A, Pleyer U, Mergler S. L-Carnitine
Suppresses Transient Receptor Potential Vanilloid Type 1 Activation in Human
Corneal Epithelial Cells. Int J Mol Sci. 2023 Jul 23;24(14):11815.

2.

Turan E, Valtink M, Reinach PS, Skupin A, Luo H, Brockmann T, Ba Salem MHO,
Pleyer U, Mergler S. L-carnitine suppresses transient receptor potential vanilloid
type 1 activity and myofibroblast transdifferentiation in human corneal
keratocytes. Lab Invest. 2021 Jun;101(6):680-689.

3.

Oronowicz J, Reinhard J, Reinach PS, Ludwiczak S, Luo H, Omar Ba Salem MH,
Kraemer MM, Biebermann H, Kakkassery V, Mergler S. Ascorbate-induced
oxidative stress mediates TRP channel activation and cytotoxicity in humanetoposide-sensitive and -resistant retinoblastoma cells. Lab Invest. 2021
Jan;101(1):70-88.

4.

Türker E, Garreis F, Khajavi N, Reinach PS, Joshi P, Brockmann T, Lucius A,
Ljubojevic N, Turan E, Cooper D, Schick F, Reinholz R, Pleyer U, Köhrle J, Mergler
S. Vascular Endothelial Growth Factor (VEGF) Induced Downstream Responses
to Transient Receptor Potential Vanilloid 1 (TRPV1) and 3-Iodothyronamine (3T1AM) in Human Corneal Keratocytes. Front Endocrinol (Lausanne). 2018 Nov
22;9:670.

5.

Walcher L, Budde C, Böhm A, Reinach PS, Dhandapani P, Ljubojevic N,
Schweiger MW, von der Waydbrink H, Reimers I, Köhrle J, Mergler S. TRPM8
Activation via 3-Iodothyronamine Blunts VEGF-Induced Transactivation of TRPV1
in Human Uveal Melanoma Cells. Front Pharmacol. 2018 Nov 13;9:1234.

6.

Bräunig J, Mergler S, Jyrch S, Hoefig CS, Rosowski M, Mittag J, Biebermann H,
Khajavi N. 3-Iodothyronamine Activates a Set of Membrane Proteins in Murine
Hypothalamic Cell Lines. Front Endocrinol (Lausanne). 2018 Sep 11;9:523.

7.

Garreis F, Schröder A, Reinach PS, Zoll S, Khajavi N, Dhandapani P, Lucius A,
Pleyer U, Paulsen F, Mergler S. Upregulation of Transient Receptor Potential
Vanilloid Type-1 Channel Activity and Ca2+ Influx Dysfunction in Human
Pterygial Cells. Invest Ophthalmol Vis Sci. 2016 May 1;57(6):2564-77.

8.

Lucius A, Khajavi N, Reinach PS, Köhrle J, Dhandapani P, Huimann P, Ljubojevic
N, Grötzinger C, Mergler S. 3-Iodothyronamine increases transient receptor
potential melastatin channel 8 (TRPM8) activity in immortalized human corneal
epithelial cells. Cell Signal. 2016 Mar;28(3):136-147.

9.

Khajavi N, Reinach PS, Slavi N, Skrzypski M, Lucius A, Strauß O, Köhrle J, Mergler S.
Thyronamine induces TRPM8 channel activation in human conjunctival epithelial
cells. Cell Signal. 2015 Feb;27(2):315-25.

antagonists such as L-carnitine may prove to be a viableapproach to suppress corneal opacification in a clinicalsetting.2
Another recent project in Mergler’s lab is devoted to thestudy of retinoblastoma, the most common type of eye
cancer in children. There remains a need to develop novel
approaches to more effectively treat this disease. Mergler’sgroup proposed that ascorbate may be regarded as apossible supportive agent in anti-cancer therapies andshowed that it acts as an oxidant, inducing tumor cell
death. The proposed mechanism involves GPCR-mediatedincreases in intracellular Ca2+ influx through TRP channels.
The scientists showed that TRP antagonists (CPZ, La3+, NAC)
inhibit ascorbate-induced currents in both etoposide-resistantand sensitive retinoblastoma cell lines.3 Very recently, theydescribed interactions between nerve growth factor (NGF)
and cannabinoid receptor 1 (CB1) that modulate control of
TRPM8 activation and, in turn, regulate sensitivity for cytostatic
drugs such as etoposide in retinoblastoma cells.21
Looking ahead, Dr. Mergler plans to continue leveraging thePort-a-Patch in both research and teaching, with exciting
projects on the horizon, such as studying pathogens including
Toxoplasma gondii.
Acknowledging the importance of the planar patch-clamp
technique for his research, he’s now giving an extra lecture
on planar patch-clamp to students in Medical Neuroscience.
“The Port-a-Patch is a very capable system that has been
instrumental in our research. Additionally, it is much easier to
use than a manual patch-clamp rig, making it an excellent
tool not only for research purposes but also for teaching andlearning patch-clamp technique,” says Dr. Mergler.

Contact Information
PD Dr. phil. nat. Stefan Mergler
Group leader, researcher, lecturer
Department of Ophthalmology
Charité – Universitätsmedizin Berlin
Corporate member of Freie Universität Berlinand Humboldt-Universität zu Berlin
Augustenburger Platz 1
13353 Berlin, Germany

Acknowledgments
We thank Stefan for sharing his views on theimportance of automated patch-clamp in his
research on the role of TRP channels in eye diseases.
We are grateful for your valuable insights andwonderful collaboration.

10. Khajavi N, Reinach PS, Skrzypski M, Lude A, Mergler S. L-carnitine reduces inhuman conjunctival epithelial cells hypertonic-induced shrinkage throughinteracting with TRPV1 channels. Cell Physiol Biochem. 2014;34(3):790-803.
11. Mergler S et al. Functional significance of thermosensitive transient receptor
potential melastatin channel 8 (TRPM8) expression in immortalized humancorneal endothelial cells. Exp Eye Res. 2013 Nov;116:337-49.
12. Mergler S et al. Calcium regulation by temperature-sensitive transient
receptor potential channels in human uveal melanoma cells. Cell Signal. 2014
Jan;26(1):56-69.
13. Yang Y, Yang H, Wang Z, Mergler S, Wolosin JM, Reinach PS. Functional TRPV1
expression in human corneal fibroblasts. Exp Eye Res. 2013 Feb;107:121-9.
14. Yang Y, Yang H, Wang Z, Varadaraj K, Kumari SS, Mergler S, Okada Y, Saika S,
Kingsley PJ, Marnett LJ, Reinach PS. Cannabinoid receptor 1 suppresses transient
receptor potential vanilloid 1-induced inflammatory responses to corneal injury.
Cell Signal. 2013 Feb;25(2):501-11.
15. Mergler S et al. Calcium regulation by thermo- and osmosensing transient
receptor potential vanilloid channels (TRPVs) in human conjunctival epithelial
cells. Histochem Cell Biol. 2012 Jun;137(6):743-61.
16. Mergler S et al. Altered calcium regulation by thermosensitive transient receptor
potential channels in etoposide-resistant WERI-Rb1 retinoblastoma cells. Exp Eye
Res. 2012 Jan;94(1):157-73.
17. Mergler S et al. Characterization of transient receptor potential vanilloid
channel 4 (TRPV4) in human corneal endothelial cells. Exp Eye Res. 2011
Nov;93(5):710-9.
18. Mergler S et al. Thermo-sensitive transient receptor potential vanilloid channel-1
regulates intracellular calcium and triggers chromogranin A secretion inpancreatic neuroendocrine BON-1 tumor cells. Cell Signal. 2012 Jan;24(1):23346.
19. Mergler S et al. Thermosensitive transient receptor potential channels in humancorneal epithelial cells. J Cell Physiol. 2011 Jul;226(7):1828-42.
20. Mergler S et al. TRPV channels mediate temperature-sensing in human corneal
endothelial cells. Exp Eye Res. 2010 Jun;90(6):758-70.
21. Ludwiczak S, Reinhard J, Reinach P.S, Li A, Oronowicz J, Yousf A, Kakkassery V,
Mergler S. Joint CB1 and NGF Receptor Activation Suppresses TRPM8 Activationin Etoposide-Resistant Retinoblastoma Cells. 2024 Jan 31;25(3):1733.

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