worked on and improved introduction

manuscript.tex

@@ -110,21 +110,31 @@ Ion channels determine neuronal excitability and mutations that alter ion channe
 \section*{Introduction} %(750 Words Maximum - Currently \textcolor{red}{837})}
 %\textit{The Introduction should briefly indicate the objectives of the study and provide enough background information to clarify why the study was undertaken and what hypotheses were tested.}
 
-Voltage-gated ion channels are vital in determining neuronal excitability, action potential generation and firing patterns \citep{bernard_channelopathies_2008, carbone_ion_2020}. In particular, the properties and combinations of ion channels and their resulting currents determine the firing properties of the neuron \citep{rutecki_neuronal_1992, pospischil_minimal_2008}. However, ion channel function can be disturbed, resulting in altered ionic current properties and altered neuronal firing behaviour\citep{carbone_ion_2020}. Ion channel mutations are a common cause of such channelopathies and are often associated with hereditary clinical disorders \citep{bernard_channelopathies_2008, carbone_ion_2020}, \textcolor{red}{such as ... }.
+Voltage-gated ion channels are vital in determining neuronal excitability, action potential generation and firing patterns \citep{bernard_channelopathies_2008, carbone_ion_2020}. In particular, the properties and combinations of ion channels and their resulting currents determine the firing properties of the neuron \citep{rutecki_neuronal_1992, pospischil_minimal_2008}. However, ion channel function can be disturbed, resulting in altered ionic current properties and altered neuronal firing behaviour\citep{carbone_ion_2020}. Ion channel mutations are a common cause of such channelopathies and are often associated with hereditary clinical disorders including ataxias, epilepsies,npain disorders, dyskinesias, intellectual disabilities, myotonias, and periodic paralyses among others \citep{bernard_channelopathies_2008, carbone_ion_2020}.
 
-Research on the effect of these altered currents on neuronal firing is important for understanding the disease and finding possible mediactions. There are a lot of widely accepted experimtental approaches. Transfection of heterologous expression systems without endogenous currents reveals changes in ion current kinetics \textcolor{red}{(cite some stuff)}. Simulations of these effects can predict their effect on neuronal firing \textcolor{red}{(cite some stuff)}. Or the influence on firing behaviour can be directly measured in transfected primary neuronal cultures \textcolor{red}{(cite some stuff)} or brain slice recrodings of mouse lines \textcolor{red}{(cite some stuff)}. 
+Although the effects of channelopathies on ion current kinetics are frequently assessed by transfection of heterologous expression systems without endogenous currents  \citep{Balestrini1044, Noebels2017, Dunlop2008} \textcolor{red}{(cite more stuff?)}, the effect of these changes in current biophysics on neuronal firing is important for understanding the pathophysiology of these disorders and for identification of potential therapeutic targets \textcolor{red}{(cite some stuff)}. Experimentally, the effects of channelopathies on neuronal firing can be assessed using primary neuronal cultures  \citep{Scalmani2006, Smith2018, Liu2019} \textcolor{red}{(cite more stuff?)} or \textit{in vitro} recordings from transgenic mouse lines \textcolor{red}{(cite some stuff)}.
 
-However, \textcolor{red}{since experimental ressources are limited,} the effect on the firing behaviour of different neuron types is often unclear \textcolor{red}{(and always incomplete)}. Different neuron types have different ion current compositions and therefore should react differently to changes in one current. In the simpler cases, the respective behaviour changes should mostly depend on the expression level of the affected current and scale with it \textcolor{red}{(cite some stuff, cite NikoPaper)}. If the change in gating kinetics is too strong, the firing behaviour can change qualitatively. Altering the relative current amplitudes in neuronal models leads to dramtic changes in their firing behaviour and dynamics \citep{rutecki_neuronal_1992, pospischil_minimal_2008,Kispersky2012, golowasch_failure_2002, barreiro_-current_2012, Noebels2017}. The same could happen for other parameters too. \citet{Liu2019} reported a drastically slowed inacitvaiton time constant for a mutation in \textcolor{red}{Na$_V$1.6}, which led to huge depolarization plateaus after an action potential, that lasted several 100 milliseconds. The most drastic example known to us would be the R1629H mutation in \textcolor{red}{SCN2A}. This mutation increases the excitability of interneurons, but decreases it in pyramidal neurons \textcolor{red}{(cite Hedrich2014 and the other paper)}. Some neuron types may be closer to certain transitions between firing states than other, making these observations even more unpredictable \textcolor{red}{(cite some bifurcation stuff?)}.
+However, experimental resources are limited the effect of a given channelopathy on different neuronal types across the brain is often unclear and not feasible to experimentally obtain. This is especially true when large numbers of distinct mutations are present and personalized medicine approaches are desired.
 
-In this study we want to get an insight into how changes in ion current kinetics change firing behaviour dependent on neuron type. We will simulate a repertoire of different neuronal models and compare their response to changes in single parameters and to changes as they were observed for mutations in \textcolor{red}{K$_V$1.1}, causing ataxia.
+%Research on the effect of these altered currents on neuronal firing is important for understanding the disease and finding possible medications. There are a lot of widely accepted experimental approaches. 
+%Transfection of heterologous expression systems without endogenous currents reveals changes in ion current kinetics \textcolor{red}{(cite some stuff)}. Simulations of these effects can predict their effect on neuronal firing \textcolor{red}{(cite some stuff)}. Or the influence on firing behaviour can be directly measured in transfected primary neuronal cultures \textcolor{red}{(cite some stuff)} or brain slice recordings of mouse lines \textcolor{red}{(cite some stuff)}. 
+%However, \textcolor{red}{since experimental resources are limited,} the effect on the firing behaviour of different neuron types is often unclear \textcolor{red}{(and always incomplete)}. 
 
+General understanding of the effects of changes in current properties on neuronal firing may help to fill the need to understand the impacts of ion channel mutations on neuronal firing. Different neuron types have different ion current compositions and therefore likely respond differently to changes in the properties of one current. For instance, altering relative current amplitudes can be dramatically alter the firing behaviour and dynamics of neurons \citep{rutecki_neuronal_1992, pospischil_minimal_2008,Kispersky2012, golowasch_failure_2002, barreiro_-current_2012, Noebels2017, Layer2021}, however other current parameters impact neuronal firing as well. \citet{Liu2019} reported that a mutation in \(\textrm{Na}_{\textrm{V}}\text{1.6}\) that drastically slowed the inactivation time constant for that channel led to large prolonged depolarization plateaus after an action potential. Another example is the R1629H SCN1A mutation which increases interneuron but decreases pyramidal neuron excitability \citep{Hedrich14874}\textcolor{red}{(and the other paper?)}. 
+The underlying dynamics of neuronal types may be different and thus may transition between firing states differently \textcolor{red}{(cite some bifurcation stuff?)}. This further increases the possible heterogeneity in firing responses to altered current properties.
+%Some neuron types may be closer to certain transitions between firing states than other, making these observations even more unpredictable \textcolor{red}{(cite some bifurcation stuff?)}.
 
-%Neuronal ion channels are vital in determining neuronal excitability, action potential generation and firing patterns \citep{bernard_channelopathies_2008, carbone_ion_2020}. In particular, the properties and combinations of ion channels and their resulting currents determine the firing properties of the neuron \citep{rutecki_neuronal_1992, pospischil_minimal_2008}. However, ion channel function can be disturb resulting in altered ionic current properties and altered neuronal firing behaviour \citep{carbone_ion_2020}. Ion channel mutations are a common cause of such channelopathies and are often associated with hereditary clinical disorders \citep{bernard_channelopathies_2008, carbone_ion_2020}. The effects of these mutations are frequently presumed \cite{Balestrini1044} or determined at a biophysical level, however assessment of the impact of mutations on neuronal firing and excitability is more difficult. Experimentally, cell culture transfection does not replicate the exact interplay of endogenous currents nor does it take into account the complexity of the nervous system including factors such as expression patterns, intracellular regulation and modulation of ion channels as well as network effects \cite{Balestrini1044, Noebels2017}. Transfected currents are characterized in isolation and the role of these isolated currents in the context of other currents in a neuron cannot be definitively inferred \cite{Dunlop2008, Noebels2017}. Additionally, transfected currents are not expressed in the presence of physiologically present auxillary proteins and are even transfected in cells of different species. Furthermore, culture conditions can shape ion channel expression \citep{ponce_expression_2018}. 
+Computational modelling approaches can be used to assess the impacts of current property changes on firing behaviour, bridging the gap between changes in the biophysical properties induced by mutations, firing and clinical symptoms. Conductance-based neuronal models enable insight into the effects of ion channel mutations with specific effects of the resulting ionic current as well as enabling \textit{in silico} assessment of the relative effects of changes in biophysical properties of ionic currents on neuronal firing. Furthermore, modelling approaches enable predictions of the effects of specific mutation and drug induced biophysical property changes. 
-% Complex interactions between different cell types and circuit level effects \textit{in vivo} are neglected in transfected cell culture. 
+
+
+%Different neuron types have different ion current compositions and therefore should react differently to changes in one current. 
+%In the simpler cases, the respective behaviour changes should mostly depend on the expression level of the affected current and scale with it \textcolor{red}{(cite some stuff, cite NikoPaper)}. If the change in gating kinetics is too strong, the firing behaviour can change qualitatively. Altering the relative current amplitudes in neuronal models leads to dramatic changes in their firing behaviour and dynamics \citep{rutecki_neuronal_1992, pospischil_minimal_2008,Kispersky2012, golowasch_failure_2002, barreiro_-current_2012, Noebels2017}. 
+%The same could happen for other parameters too. \citet{Liu2019} reported a drastically slowed inactivation time constant for a mutation in \textcolor{red}{Na$_V$1.6}, which led to huge depolarization plateaus after an action potential, that lasted several 100 milliseconds. The most drastic example known to us would be the R1629H mutation in \textcolor{red}{SCN2A}. This mutation increases the excitability of interneurons, but decreases it in pyramidal neurons \textcolor{red}{(cite Hedrich2014 and the other paper)}. Some neuron types may be closer to certain transitions between firing states than other, making these observations even more unpredictable \textcolor{red}{(cite some bifurcation stuff?)}.
+
+We therefore investigate the role that neuronal type plays on the outcome of ion current kinetic changes on firing by simulating the response of a repertoire of different neuronal models to changes in single current parameters as well as to episodic ataxia type 1 associated \Kv mutations.
+
+%In this study we want to get an insight into how changes in ion current kinetics change firing behaviour dependent on neuron type. We will simulate a repertoire of different neuronal models and compare their response to changes in single parameters and to changes as they were observed for mutations in \Kv, causing ataxia.
 
-%Ion channel transfection of primary neuronal cultures can overcome some of the limitations of cell culture expression. In transfected neuronal cell cultures firing can more readily be assessed as endogenous currents are present, however the expressed and endogenous versions of the same ion channel are present in the cell \cite{Scalmani2006, Smith2018}. To avoid the confound of both expressed and endogenous current contributing to firing, a drug resistance can be introduced to the transfected ion channel and  the endogenous version of this current can be pharmacologically silenced \cite{Liu2019}.  Although addition of TTX-resistance to \(\textrm{Na}_{\textrm{V}}\) does not alter the gating properties of these channels \cite{Leffler2005}, the relative expression of the transfected ion channel in relation to endogenous currents can be variable and non-specific blocking of ion channels not affected by the channelopathy may occur. As the firing behaviour and dynamics of neuronal models can be dramatically altered by altering relative current amplitudes \citep{rutecki_neuronal_1992, pospischil_minimal_2008,Kispersky2012, golowasch_failure_2002, barreiro_-current_2012, Noebels2017}, primary neuronal cultures provide a useful general indication as to the effects of ion channel mutations but do not provide definitive insight into the effects of a channelopathy on \textit{in vivo} firing. 
-   
-%The generation of mice lines is costly and behavioural characterization of new mice lines is required to assess similarities to patient symptoms. Although the generation of mouse lines is desirable for a clinical disorder characterized by a specific ion channel mutation, this approach becomes impractical for personalized treatment for large numbers of distinct mutations. General understanding of the effects of changes in current properties on neuronal firing may help to fill the need to understand the impacts of ion channel mutations on neuronal firing. Specifically, modelling approaches can be used to assess the impacts of current property changes on firing behaviour, bridging the gap between changes in the biophysical properties induced by mutations and clinical symptoms. Conductance-based neuronal models enable insight into the effects of ion channel mutations with specific effects of the resulting ionic current as well as enabling \textit{in silico} assessment of the relative effects of changes in biophysical properties of ionic currents on neuronal firing. The effects of altered voltage-gated potassium channel \Kv function is of particular interest in this study as it gives rise to the \IKv current and is associated with episodic ataxia type 1. Furthermore, modelling approaches enable predictions of the effects of specific mutation and drug induced biophysical property changes.
 
 %\Kv channels, encoded by the KCNA1 gene, play a role in repolarizing the action potential, neuronal firing patterns, neurotransmitter release, and saltatory conduction \citep{dadamo_episodic_1998} and are expressed throughout the CNS  \citep{tsaur_differential_1992, wang_localization_1994, veh_immunohistochemical_1995}.   
 %Altered \Kv channel function as a result of KCNA1 mutations in humans is associated with episodic ataxia type 1 (EA1) which is characterized by period attacks of ataxia and persistent myokymia \citep{parker_periodic_1946, van_dyke_hereditary_1975}. 
@@ -133,6 +143,19 @@ In this study we want to get an insight into how changes in ion current kinetics
 
 %Using a diverse set of conductance-based neuronal models we examine the role of current environment on the impact of alterations in channels properties on firing behavior generally and for EA1 associated \Kv mutations. 
 
+%In this study we want to get an insight into how changes in ion current kinetics change firing behaviour dependent on neuron type. We will simulate a repertoire of different neuronal models and compare their response to changes in single parameters and to changes as they were observed for mutations in \textcolor{red}{K$_V$1.1}, causing ataxia.
+
+
+%Neuronal ion channels are vital in determining neuronal excitability, action potential generation and firing patterns \citep{bernard_channelopathies_2008, carbone_ion_2020}. In particular, the properties and combinations of ion channels and their resulting currents determine the firing properties of the neuron \citep{rutecki_neuronal_1992, pospischil_minimal_2008}. However, ion channel function can be disturb resulting in altered ionic current properties and altered neuronal firing behaviour \citep{carbone_ion_2020}. Ion channel mutations are a common cause of such channelopathies and are often associated with hereditary clinical disorders \citep{bernard_channelopathies_2008, carbone_ion_2020}. The effects of these mutations are frequently presumed \cite{Balestrini1044} or determined at a biophysical level, however assessment of the impact of mutations on neuronal firing and excitability is more difficult. 
+
+%Experimentally, cell culture transfection does not replicate the exact interplay of endogenous currents nor does it take into account the complexity of the nervous system including factors such as expression patterns, intracellular regulation and modulation of ion channels as well as network effects \cite{Balestrini1044, Noebels2017}. Transfected currents are characterized in isolation and the role of these isolated currents in the context of other currents in a neuron cannot be definitively inferred \cite{Dunlop2008, Noebels2017}. Additionally, transfected currents are not expressed in the presence of physiologically present auxillary proteins and are even transfected in cells of different species. Furthermore, culture conditions can shape ion channel expression \citep{ponce_expression_2018}. 
+% Complex interactions between different cell types and circuit level effects \textit{in vivo} are neglected in transfected cell culture. 
+
+%Ion channel transfection of primary neuronal cultures can overcome some of the limitations of cell culture expression. In transfected neuronal cell cultures firing can more readily be assessed as endogenous currents are present, however the expressed and endogenous versions of the same ion channel are present in the cell \cite{Scalmani2006, Smith2018}. To avoid the confound of both expressed and endogenous current contributing to firing, a drug resistance can be introduced to the transfected ion channel and  the endogenous version of this current can be pharmacologically silenced \cite{Liu2019}.  Although addition of TTX-resistance to \(\textrm{Na}_{\textrm{V}}\) does not alter the gating properties of these channels \cite{Leffler2005}, the relative expression of the transfected ion channel in relation to endogenous currents can be variable and non-specific blocking of ion channels not affected by the channelopathy may occur. As the firing behaviour and dynamics of neuronal models can be dramatically altered by altering relative current amplitudes \citep{rutecki_neuronal_1992, pospischil_minimal_2008,Kispersky2012, golowasch_failure_2002, barreiro_-current_2012, Noebels2017}, primary neuronal cultures provide a useful general indication as to the effects of ion channel mutations but do not provide definitive insight into the effects of a channelopathy on \textit{in vivo} firing. 
+   
+%The generation of mice lines is costly and behavioural characterization of new mice lines is required to assess similarities to patient symptoms. Although the generation of mouse lines is desirable for a clinical disorder characterized by a specific ion channel mutation, this approach becomes impractical for personalized treatment for large numbers of distinct mutations. General understanding of the effects of changes in current properties on neuronal firing may help to fill the need to understand the impacts of ion channel mutations on neuronal firing. Specifically, modelling approaches can be used to assess the impacts of current property changes on firing behaviour, bridging the gap between changes in the biophysical properties induced by mutations and clinical symptoms. Conductance-based neuronal models enable insight into the effects of ion channel mutations with specific effects of the resulting ionic current as well as enabling \textit{in silico} assessment of the relative effects of changes in biophysical properties of ionic currents on neuronal firing. The effects of altered voltage-gated potassium channel \Kv function is of particular interest in this study as it gives rise to the \IKv current and is associated with episodic ataxia type 1. Furthermore, modelling approaches enable predictions of the effects of specific mutation and drug induced biophysical property changes.
+
+
 
 \par\null
 

ref.bib

@@ -1377,4 +1377,18 @@ SIGNIFICANCE: Bromide is most effective and is a well-tolerated drug among DS pa
   urldate  = {2022-04-12},
 }
 
+ 
+@Article{Layer2021,
+  author   = {Layer, Nikolas and Sonnenberg, Lukas and Pardo González, Emilio and Benda, Jan and Hedrich, Ulrike B. S. and Lerche, Holger and Koch, Henner and Wuttke, Thomas V.},
+  journal  = {Frontiers in Cellular Neuroscience},
+  title    = {Dravet {Variant} {SCN1AA1783V} {Impairs} {Interneuron} {Firing} {Predominantly} by {Altered} {Channel} {Activation}},
+  year     = {2021},
+  issn     = {1662-5102},
+  volume   = {15},
+  abstract = {Dravet syndrome (DS) is a developmental epileptic encephalopathy mainly caused by functional NaV1.1 haploinsufficiency in inhibitory interneurons. Recently, a new conditional mouse model expressing the recurrent human p.(Ala1783Val) missense variant has become available. In this study, we provided an electrophysiological characterization of this variant in tsA201 cells, revealing both altered voltage-dependence of activation and slow inactivation without reduced sodium peak current density. Based on these data, simulated interneuron (IN) firing properties in a conductance-based single-compartment model suggested surprisingly similar firing deficits for NaV1.1A1783V and full haploinsufficiency as caused by heterozygous truncation variants. Impaired NaV1.1A1783V channel activation was predicted to have a significantly larger impact on channel function than altered slow inactivation and is therefore proposed as the main mechanism underlying IN dysfunction. The computational model was validated in cortical organotypic slice cultures derived from conditional Scn1aA1783V mice. Pan-neuronal activation of the p.Ala1783V in vitro confirmed a predicted IN firing deficit and revealed an accompanying reduction of interneuronal input resistance while demonstrating normal excitability of pyramidal neurons. Altered input resistance was fed back into the model for further refinement. Taken together these data demonstrate that primary loss of function (LOF) gating properties accompanied by altered membrane characteristics may match effects of full haploinsufficiency on the neuronal level despite maintaining physiological peak current density, thereby causing DS.},
+  file     = {Full Text PDF:https\://www.frontiersin.org/articles/10.3389/fncel.2021.754530/pdf:application/pdf},
+  url      = {https://www.frontiersin.org/article/10.3389/fncel.2021.754530},
+  urldate  = {2022-04-27},
+}
+
 @Comment{jabref-meta: databaseType:bibtex;}