Primoz Pirih | The Graduate University for Advanced Studies (original) (raw)
Papers by Primoz Pirih
Frontiers in Physiology, 2013
Lecture Notes in Computer Science, 2014
ABSTRACT
Journal of Neuroscience, 2007
Postsynaptic recordings from this synapse in prehearing animals had delivered strong indications ... more Postsynaptic recordings from this synapse in prehearing animals had delivered strong indications for synchronized release of several vesicles. The underlying mechanism, however, remains unclear. Here, we used presynaptic membrane capacitance measurements to test whether IHCs release vesicles in a statistically independent or dependent (coordinated) manner. Exocytic changes of membrane capacitance (⌬C m ) were repeatedly stimulated in IHCs of prehearing and hearing mice by short depolarizations to preferentially recruit the readily releasable pool of synaptic vesicles. A compound Poisson model was devised to describe hair cell exocytosis and to test the analysis. From the trial-to-trial fluctuations of the ⌬C m we were able to estimate the apparent size of the elementary fusion event (C app ) at the hair cell synapse to be 96 -223 aF in immature and 55-149 aF in mature IHCs. We also approximated the single vesicle capacitance in IHCs by measurements of synaptic vesicle diameters in electron micrographs. The results (immature, 48 aF; mature, 45 aF) were lower than the respective C app estimates. This indicates that coordinated exocytosis of synaptic vesicles occurs at both immature and mature hair cell synapses. Approximately 35% of the release events in mature IHCs and ϳ50% in immature IHCs were predicted to involve coordinated fusion, when assuming a geometric distribution of elementary sizes. In summary, our presynaptic measurements indicate coordinated exocytosis but argue for a lesser degree of coordination than suggested by postsynaptic recordings.
Journal of Experimental Biology, 2013
The owlfly Libelloides macaronius (Insecta: Neuroptera) has large bipartite eyes of the superposi... more The owlfly Libelloides macaronius (Insecta: Neuroptera) has large bipartite eyes of the superposition type. The spatial resolution and sensitivity of the photoreceptor array in the dorsofrontal eye part was studied with optical and electrophysiological methods. Using structured illumination microscopy, the interommatidial angle in the central part of the dorsofrontal eye was determined to be Δj=1.1deg. Eye shine measurements with an epi-illumination microscope yielded an effective superposition pupil size of about 300 facets. Intracellular recordings confirmed that all photoreceptors were UV-receptors (λ max =350nm). The average photoreceptor acceptance angle was 1.8deg, with a minimum of 1.4deg. The receptor dynamic range was two log units, and the Hill coefficient of the intensity-response function was n=1.2. The signal-to-noise ratio of the receptor potential was remarkably high and constant across the whole dynamic range (root mean square r.m.s. noise=0.5% V max ). Quantum bumps could not be observed at any light intensity, indicating low voltage gain. Presumably, the combination of large aperture superposition optics feeding an achromatic array of relatively insensitive receptors with a steep intensity-response function creates a low-noise, high spatial acuity instrument. The sensitivity shift to the UV range reduces the clutter created by clouds within the sky image. These properties of the visual system are optimal for detecting small insect prey as contrasting spots against both clear and cloudy skies.
Beta cells of pancreatic islets are blood glucose level sensors and effectors of insulin release.... more Beta cells of pancreatic islets are blood glucose level sensors and effectors of insulin release. One of the possible key players in metabolism-membrane excitation coupling, which leads to exocytosis of hormone-filled vesicles, are ATP-sensitive K + channels. There are strong objections against the prevailing role of these channels in coupling because of their almost complete inactivation at physiological ATP level. We measured macroscopic currents in whole-cell patched isolated beta cells and tried to assess whether the changes in these channels' activity upon ATP depletion with dialysis could be detected with stationary and non-stationary analysis of current noise.
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Toxicology in Vitro, 2008
Corrigendum to ''Detection of xenobiotic-induced DNA damage by the comet assay applied to human a... more Corrigendum to ''Detection of xenobiotic-induced DNA damage by the comet assay applied to human and rat precision-cut liver slices" [
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2011
The males of many pierid butterflies have iridescent wings, which presumably function in intraspe... more The males of many pierid butterflies have iridescent wings, which presumably function in intraspecific communication. The iridescence is due to nanostructured ridges of the cover scales. We have studied the iridescence in the males of a few members of Coliadinae, Gonepteryx aspasia, G. cleopatra, G. rhamni, and Colias croceus, and in two members of the Colotis group, Hebomoia glaucippe and Colotis regina. Imaging scatterometry demonstrated that the pigmentary colouration is diffuse whereas the structural colouration creates a directional, line-shaped far-field radiation pattern. Angle-dependent reflectance measurements demonstrated that the directional iridescence distinctly varies among closely related species. The species-dependent scale curvature determines the spatial properties of the wing iridescence. Narrow beam illumination of flat scales results in a narrow far-field iridescence pattern, but curved scales produce broadened patterns. The restricted spatial visibility of iridescence presumably plays a role in intraspecific signalling.
Toxicology in Vitro, 2007
The comet assay is a simple and sensitive method for measuring DNA damage at the level of individ... more The comet assay is a simple and sensitive method for measuring DNA damage at the level of individual cells and is extensively used in genotoxicity studies. It is commonly applied to cultured cells. The aim of this study was to apply the comet assay for use in fresh liver tissue, where metabolic activity, all cell types and tissue architecture are preserved.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2010
We have simultaneously measured the electroretinogram (ERG) and the metarhodopsin content via flu... more We have simultaneously measured the electroretinogram (ERG) and the metarhodopsin content via fluorescence in white-eyed, wild-type Drosophila and the arrestin2 hypomorphic mutant (w − ;arr2 3 ) at a range of stimulus wavelengths and intensities. Photoreceptor response amplitude and termination (transition between full repolarization and prolonged depolarizing afterpotential, PDA) were related to visual pigment conversions and arrestin concentration. The data were implemented in a kinetic model of the rhodopsin–arrestin cycle, allowing us to estimate the active metarhodopsin concentration as a function of effective light intensity and arrestin concentration. Arrestin reduction in the mutant modestly increased the light sensitivity and decreased the photoreceptor dynamic range. Compared to the wild type, in the mutant the transition between full repolarization and PDA occurred at a lower metarhodopsin fraction and was more abrupt. We developed a steady-state stochastic model to interpret the dependence of the PDA on effective light intensity and arrestin content and to help deduce the arrestin to rhodopsin ratio from the sensitivity and PDA data. The feasibility of different experimental methods for the estimation of arrestin content from ERG and PDA is discussed.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2010
We studied the spectral and polarisation sensitivities of photoreceptors of the butterfly Colias ... more We studied the spectral and polarisation sensitivities of photoreceptors of the butterfly Colias erate by using intracellular electrophysiological recordings and stimulation with light pulses. We developed a method of response waveform comparison (RWC) for evaluating the effective intensity of the light pulses. We identified one UV, four violet-blue, two green and two red photoreceptor classes. We estimated the peak wavelengths of four rhodopsins to be at about 360, 420, 460 and 560 nm. The four violet-blue classes are presumably based on combinations of two rhodopsins and a violet-absorbing screening pigment. The green classes have reduced sensitivity in the ultraviolet range. The two red classes have primary peaks at about 650 and 665 nm, respectively, and secondary peaks at about 480 nm. The shift of the main peak, so far the largest amongst insects, is presumably achieved by tuning the effective thickness of the red perirhabdomal screening pigment. Polarisation sensitivity of green and red photoreceptors is higher at the secondary than at the main peak. We found a 20-fold variation of sensitivity within the cells of one green class, implying possible photoreceptor subfunctionalisation. We propose an allocation scheme of the receptor classes into the three ventral ommatidial types.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2011
The wings of most pierid butterflies exhibit a main, pigmentary colouration: white, yellow or ora... more The wings of most pierid butterflies exhibit a main, pigmentary colouration: white, yellow or orange. The males of many species have in restricted areas of the wing upper sides a distinct structural colouration, which is created by stacks of lamellae in the ridges of the wing scales, resulting in iridescence. The amplitude of the reflectance is proportional to the number of lamellae in the ridge stacks. The angle-dependent peak wavelength of the observed iridescence is in agreement with classical multilayer theory. The iridescence is virtually always in the ultraviolet wavelength range, but some species have a blue-peaking iridescence. The spectral properties of the pigmentary and structural colourations are presumably tuned to the spectral sensitivities of the butterflies’ photoreceptors.
Journal of Experimental Biology, 2009
Here we report the remarkable anatomy of the eye of the Eastern Pale Clouded yellow butterfly, Co... more Here we report the remarkable anatomy of the eye of the Eastern Pale Clouded yellow butterfly, Colias erate. An ommatidium of C. erate bears nine photoreceptors, R1-9, which together form a tiered and fused rhabdom. The distal tier of the rhabdom consists of the rhabdomeral microvilli of R1-4 photoreceptors, R5-8 photoreceptors contribute the proximal tier, and the R9 photoreceptor adds a few microvilli at the base. In transverse sections, four spots of red pigment surrounding the rhabdom are evident in the ventral region of the eye. The red pigment acts as a strong red filter for the proximal photoreceptors. The arrangement of the pigment spots distinguishes the ommatidia into three types: trapezoidal (type I), square (type II) and rectangular (type III). In all types of ommatidia, the distal and the proximal tiers of the rhabdom are divided by a strong constriction, clearly to enhance the filtering effect of the red pigment. The ommatidial heterogeneity can also be observed by optical measurements. The eye shine, resulting from tapetal reflections, peaks in type I ommatidia at 660 nm, and in type II and III ommatidia at 730 nm. The far-red peaking eye shine indicates that C. erate has far-red-sensitive photoreceptors. Type I ommatidia fluoresce under violet excitation, implying the presence of a violet-absorbing pigment that acts as a short-wavelength filter.
Frontiers in Physiology, 2013
Lecture Notes in Computer Science, 2014
ABSTRACT
Journal of Neuroscience, 2007
Postsynaptic recordings from this synapse in prehearing animals had delivered strong indications ... more Postsynaptic recordings from this synapse in prehearing animals had delivered strong indications for synchronized release of several vesicles. The underlying mechanism, however, remains unclear. Here, we used presynaptic membrane capacitance measurements to test whether IHCs release vesicles in a statistically independent or dependent (coordinated) manner. Exocytic changes of membrane capacitance (⌬C m ) were repeatedly stimulated in IHCs of prehearing and hearing mice by short depolarizations to preferentially recruit the readily releasable pool of synaptic vesicles. A compound Poisson model was devised to describe hair cell exocytosis and to test the analysis. From the trial-to-trial fluctuations of the ⌬C m we were able to estimate the apparent size of the elementary fusion event (C app ) at the hair cell synapse to be 96 -223 aF in immature and 55-149 aF in mature IHCs. We also approximated the single vesicle capacitance in IHCs by measurements of synaptic vesicle diameters in electron micrographs. The results (immature, 48 aF; mature, 45 aF) were lower than the respective C app estimates. This indicates that coordinated exocytosis of synaptic vesicles occurs at both immature and mature hair cell synapses. Approximately 35% of the release events in mature IHCs and ϳ50% in immature IHCs were predicted to involve coordinated fusion, when assuming a geometric distribution of elementary sizes. In summary, our presynaptic measurements indicate coordinated exocytosis but argue for a lesser degree of coordination than suggested by postsynaptic recordings.
Journal of Experimental Biology, 2013
The owlfly Libelloides macaronius (Insecta: Neuroptera) has large bipartite eyes of the superposi... more The owlfly Libelloides macaronius (Insecta: Neuroptera) has large bipartite eyes of the superposition type. The spatial resolution and sensitivity of the photoreceptor array in the dorsofrontal eye part was studied with optical and electrophysiological methods. Using structured illumination microscopy, the interommatidial angle in the central part of the dorsofrontal eye was determined to be Δj=1.1deg. Eye shine measurements with an epi-illumination microscope yielded an effective superposition pupil size of about 300 facets. Intracellular recordings confirmed that all photoreceptors were UV-receptors (λ max =350nm). The average photoreceptor acceptance angle was 1.8deg, with a minimum of 1.4deg. The receptor dynamic range was two log units, and the Hill coefficient of the intensity-response function was n=1.2. The signal-to-noise ratio of the receptor potential was remarkably high and constant across the whole dynamic range (root mean square r.m.s. noise=0.5% V max ). Quantum bumps could not be observed at any light intensity, indicating low voltage gain. Presumably, the combination of large aperture superposition optics feeding an achromatic array of relatively insensitive receptors with a steep intensity-response function creates a low-noise, high spatial acuity instrument. The sensitivity shift to the UV range reduces the clutter created by clouds within the sky image. These properties of the visual system are optimal for detecting small insect prey as contrasting spots against both clear and cloudy skies.
Beta cells of pancreatic islets are blood glucose level sensors and effectors of insulin release.... more Beta cells of pancreatic islets are blood glucose level sensors and effectors of insulin release. One of the possible key players in metabolism-membrane excitation coupling, which leads to exocytosis of hormone-filled vesicles, are ATP-sensitive K + channels. There are strong objections against the prevailing role of these channels in coupling because of their almost complete inactivation at physiological ATP level. We measured macroscopic currents in whole-cell patched isolated beta cells and tried to assess whether the changes in these channels' activity upon ATP depletion with dialysis could be detected with stationary and non-stationary analysis of current noise.
[
Toxicology in Vitro, 2008
Corrigendum to ''Detection of xenobiotic-induced DNA damage by the comet assay applied to human a... more Corrigendum to ''Detection of xenobiotic-induced DNA damage by the comet assay applied to human and rat precision-cut liver slices" [
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2011
The males of many pierid butterflies have iridescent wings, which presumably function in intraspe... more The males of many pierid butterflies have iridescent wings, which presumably function in intraspecific communication. The iridescence is due to nanostructured ridges of the cover scales. We have studied the iridescence in the males of a few members of Coliadinae, Gonepteryx aspasia, G. cleopatra, G. rhamni, and Colias croceus, and in two members of the Colotis group, Hebomoia glaucippe and Colotis regina. Imaging scatterometry demonstrated that the pigmentary colouration is diffuse whereas the structural colouration creates a directional, line-shaped far-field radiation pattern. Angle-dependent reflectance measurements demonstrated that the directional iridescence distinctly varies among closely related species. The species-dependent scale curvature determines the spatial properties of the wing iridescence. Narrow beam illumination of flat scales results in a narrow far-field iridescence pattern, but curved scales produce broadened patterns. The restricted spatial visibility of iridescence presumably plays a role in intraspecific signalling.
Toxicology in Vitro, 2007
The comet assay is a simple and sensitive method for measuring DNA damage at the level of individ... more The comet assay is a simple and sensitive method for measuring DNA damage at the level of individual cells and is extensively used in genotoxicity studies. It is commonly applied to cultured cells. The aim of this study was to apply the comet assay for use in fresh liver tissue, where metabolic activity, all cell types and tissue architecture are preserved.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2010
We have simultaneously measured the electroretinogram (ERG) and the metarhodopsin content via flu... more We have simultaneously measured the electroretinogram (ERG) and the metarhodopsin content via fluorescence in white-eyed, wild-type Drosophila and the arrestin2 hypomorphic mutant (w − ;arr2 3 ) at a range of stimulus wavelengths and intensities. Photoreceptor response amplitude and termination (transition between full repolarization and prolonged depolarizing afterpotential, PDA) were related to visual pigment conversions and arrestin concentration. The data were implemented in a kinetic model of the rhodopsin–arrestin cycle, allowing us to estimate the active metarhodopsin concentration as a function of effective light intensity and arrestin concentration. Arrestin reduction in the mutant modestly increased the light sensitivity and decreased the photoreceptor dynamic range. Compared to the wild type, in the mutant the transition between full repolarization and PDA occurred at a lower metarhodopsin fraction and was more abrupt. We developed a steady-state stochastic model to interpret the dependence of the PDA on effective light intensity and arrestin content and to help deduce the arrestin to rhodopsin ratio from the sensitivity and PDA data. The feasibility of different experimental methods for the estimation of arrestin content from ERG and PDA is discussed.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2010
We studied the spectral and polarisation sensitivities of photoreceptors of the butterfly Colias ... more We studied the spectral and polarisation sensitivities of photoreceptors of the butterfly Colias erate by using intracellular electrophysiological recordings and stimulation with light pulses. We developed a method of response waveform comparison (RWC) for evaluating the effective intensity of the light pulses. We identified one UV, four violet-blue, two green and two red photoreceptor classes. We estimated the peak wavelengths of four rhodopsins to be at about 360, 420, 460 and 560 nm. The four violet-blue classes are presumably based on combinations of two rhodopsins and a violet-absorbing screening pigment. The green classes have reduced sensitivity in the ultraviolet range. The two red classes have primary peaks at about 650 and 665 nm, respectively, and secondary peaks at about 480 nm. The shift of the main peak, so far the largest amongst insects, is presumably achieved by tuning the effective thickness of the red perirhabdomal screening pigment. Polarisation sensitivity of green and red photoreceptors is higher at the secondary than at the main peak. We found a 20-fold variation of sensitivity within the cells of one green class, implying possible photoreceptor subfunctionalisation. We propose an allocation scheme of the receptor classes into the three ventral ommatidial types.
Journal of Comparative Physiology A-neuroethology Sensory Neural and Behavioral Physiology, 2011
The wings of most pierid butterflies exhibit a main, pigmentary colouration: white, yellow or ora... more The wings of most pierid butterflies exhibit a main, pigmentary colouration: white, yellow or orange. The males of many species have in restricted areas of the wing upper sides a distinct structural colouration, which is created by stacks of lamellae in the ridges of the wing scales, resulting in iridescence. The amplitude of the reflectance is proportional to the number of lamellae in the ridge stacks. The angle-dependent peak wavelength of the observed iridescence is in agreement with classical multilayer theory. The iridescence is virtually always in the ultraviolet wavelength range, but some species have a blue-peaking iridescence. The spectral properties of the pigmentary and structural colourations are presumably tuned to the spectral sensitivities of the butterflies’ photoreceptors.
Journal of Experimental Biology, 2009
Here we report the remarkable anatomy of the eye of the Eastern Pale Clouded yellow butterfly, Co... more Here we report the remarkable anatomy of the eye of the Eastern Pale Clouded yellow butterfly, Colias erate. An ommatidium of C. erate bears nine photoreceptors, R1-9, which together form a tiered and fused rhabdom. The distal tier of the rhabdom consists of the rhabdomeral microvilli of R1-4 photoreceptors, R5-8 photoreceptors contribute the proximal tier, and the R9 photoreceptor adds a few microvilli at the base. In transverse sections, four spots of red pigment surrounding the rhabdom are evident in the ventral region of the eye. The red pigment acts as a strong red filter for the proximal photoreceptors. The arrangement of the pigment spots distinguishes the ommatidia into three types: trapezoidal (type I), square (type II) and rectangular (type III). In all types of ommatidia, the distal and the proximal tiers of the rhabdom are divided by a strong constriction, clearly to enhance the filtering effect of the red pigment. The ommatidial heterogeneity can also be observed by optical measurements. The eye shine, resulting from tapetal reflections, peaks in type I ommatidia at 660 nm, and in type II and III ommatidia at 730 nm. The far-red peaking eye shine indicates that C. erate has far-red-sensitive photoreceptors. Type I ommatidia fluoresce under violet excitation, implying the presence of a violet-absorbing pigment that acts as a short-wavelength filter.