Stereology for Biological Research with a Focus on Neuroscience
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We guide our gaze to what we want to look at; we choose and approach what to touch smell or taste. In addition our motor actions are carried out in the light of sensory inputs. Our research focuses on understanding how sensory inputs are integrated with motor actions to produce a unified percept.
We specifically investigate sensorimotor circuitry of somatosensation during active exploratory behavior in head-restrained mobile mice. We use 2-photon calcium imaging to measure neuronal activity and benefit from viral and genetic constructs to dissect and manipulate the circuitry. Keywords: sensory processing, locomotion, in vivo physiology, optical imaging, sensorimotor integration, neural circuits.
Dominik R. Our knowledge on the molecular and neuronal basis of defensive emotions such as panic, learned fear, and anxiety, mainly comes from animal research.
Yet, animal models of emotion are not directly comparable to human emotion. The challenge that our lab seeks to address is to translate established animal models of emotion to humans, by creating and investigating behavioural cross-species test beds. We are particularly interested in a computational formulation of aversive learning fear conditioning , anxiety, and other defensive behaviours.
To this end, we use structural and functional MRI, MEG, peripheral physiology, behavioural experiments, and computational modelling. Keywords : Emotions, comparative neuroscience, computational modelling, fear conditioning, anxiety. Publications : pubmed. Research Focus : Our research focuses on a group of human Mendelian disorders called ciliopathies which are unified by shared genetic causes resulting in primary cilium dysfunction. Primary cilia are small non-motile organelles present on the surface of most vertebrate cells where they are involved in transduction of sensory, mechanical or chemical signals and in regulation of signaling pathways during development and cell homeostasis.
Typical clinical presentations of ciliopathies include neurological involvement, retinal degeneration and renal fibrocystic disease, as illustrated by Joubert syndrome JS , an iconic ciliopathy which is the main focus of our research. Keywords : ciliopathies, primary cilia, Joubert syndrome, zebrafish, iPSC. Department of Neurology, University Hospital Zurich christian. Our research aims to understand the development of tissue-specific autoimmunity in particular in the context of interactions of the nervous system with the immune system. Our main research interests can be categorized as such:.
Institute of Pharmacology and Toxicology, University of Zurich benke pharma. Keywords: stereology, optical fractionator, optical disector, cavalieri estimator, neuronal injury. Introduction Neuronal injury and neurodegeneration are hallmark pathologies of a variety of acute and chronic neurological conditions such as traumatic brain injury, stroke and epilepsy.
Open in a separate window. Figure 1. NeuN Immunohistochemistry The procedure employed for NeuN immunohistochemistry was based on previous report in Rao et al. PV Immunohistochemistry The procedure employed for PV immunohistochemistry was outlined in previous report in Kuruba et al. Stereology Imaging and Drawing Super images of the entire slide containing hippocampal sections three slices in each slide were acquired by stereology newCAST software Visiopharm, Denmark with a 1.
Table 1 Step process for stereology. Turn on Microscope 2. Load slide and select and focus the 1. Click Finish B. Repeat steps until every ROI is accounted for D. Click Finish and save data Repeat until all ROI have been accounted for F. Repeat for all ROI. Figure 3. Figure 2. Figure 6. Figure 4. Table 2 Counting references for stereological analysis of neuronal tissue volume and cell density for NeuN and parvalbumin immunostaining in rat. Optical Fractionator and Sampling Scheme Method The optical fractionator combines the optical disector, a 3D probe used for counting neurons, and the fractionator, a systematic uniform sampling scheme of a known fraction of the region being analyzed.
The Optical Disector The optical disector consists of an unbiased counting frame that is moved through a relatively thick portion of tissue by focusing the microscope. Stereological Parameters for Mice Even though rats have been widely used for studying neuronal diseases, mice are used more frequently for studies involving genetic modeling, neuronal plasticity, and neurodegeneration. Figure 5.
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Quantification of Principal Neurons, Interneurons and Total Neurons in the Hippocampus Subfields To validate the stereology protocol, the absolute number of total cells and interneurons were separately quantified in each hippocampus subfield in rat brain sections stained with NeuN or PV.
Figure 7. Figure 8. Discussion of Main Protocol The present study shows successful optimization of a neurostereology technique for absolute quantification of specific types of neurons and interneurons in the brain sections. Advantages of the Method One of the advantages of the current method is that it combines several elements of similar stereological methods to create a more efficient process.
Frontiers | Editorial: Neurostereology | Frontiers in Neuroanatomy
Potential Problem Areas and Solutions One of the most difficult challenges of stereology is defining the top and bottom positions of the slice under the microscope Geuna and Herrera-Rincon, Conclusion In conclusion, these results suggest that the optimized neurostereology protocol is a powerful tool for studies of neuronal injury and neurodegeneration by absolute quantification of total neurons and interneurons in the brain. Conflict of Interest Statement The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.
References Abercrombie M. Estimation of nuclear population from microtome sections. Quantitative analysis of GABA-like-immunoreactive and parvalbumin-containing neurons in the CA1 region of the rat hippocampus using a stereological method, the disector. Brain Res. Estimation of surface area from vertical sections. Design-based stereology: introduction to basic concepts and practical approaches for estimation of cell number. The impact of recent stereological advances on quantitative studies of the nervous system. Methods 18 , 39— Effects of low-intensity pulsed ultrasound on healing of mandibular bone defects: an experimental study in rabbits.
Estimating cell number in the central nervous system by stereological methods: the optical disector and fractionator. Sampling theory and automated simulations for vertical sections, applied to human brain.
Sampling designs for stereology. Stereology of single objects. Precision of the fractionator from cavalieri designs. Stereological approaches to identifying neuropathology in psychosis. Psychiatry 69 , — Tissue shrinkage and unbiased stereological estimation of particle number and size. Untersuchungen uber den Bau der menschlichen Hypophyse mit besonderer Berucksichtigung der quantitative mikromorphologischen Verhaltnisse. Update on stereology for light microscopy. Cell Tissue Res.
The new stereological tools: disector, fractionator, nucleator and point sampled intercepts and their use in pathological research and diagnosis.
Basic Application of Stereology in Histology and Medical Sciences
APMIS 96 , — The efficiency of systematic sampling in stereology—reconsidered. Notes on the estimation of the numerical density of arbitrary profiles: the edge effect. Practical considerations for the use of the optical disector in estimating neuronal number.
Methods 51 , 83— Acute seizures in old age leads to a greater loss of CA1 pyramidal neurons, an increased propensity for developing chronic TLE and a severe cognitive dysfunction. Aging Dis. Fractional volume of placental vessels in women with diabetes using a novel stereological 3D power Doppler technique.
Placenta 34 , — The disector counting technique. NeuroQuantology 10 , 44— How to use the optical fractionator: an example based on the estimation of neurons in the hippocampal CA1 and CA3 regions of tree shrews. Differential susceptibility of interneurons expressing neuropeptide y or parvalbumin in the aged hippocampus to acute seizure activity.
PLoS One 6 :e Benzodiazepine resistant status epilepticus and brain injury in DFP model of organophosphate intoxication. Epilepsy Soc. Quantitative immunocytochemistry at the ultrastructural level: a stereology-based approach to molecular nanomorphomics. Laminar distribution of non-principal neurons in the rat hippocampus, with special reference to their compositional difference among layers.
Total number of neurons and glial cells in human brain nuclei estimated by the disector and the fractionator. The Rat Brain in Stereotaxic Coordinates. New York: Academic Press. Hippocampal neurodegeneration, spontaneous seizures and mossy fiber sprouting in the F rat model of temporal lobe epilepsy. Counting sectioned cells via mathematical reconstruction. Stereology: a method for analyzing images. The efficiency of design-based stereology in estimating spiral ganglion populations in mice.
Stereologic analysis of the lateral geniculate nucleus of the thalamus in normal and schizophrenic subjects. Psychiatry Res. No further loss of dorsal root ganglion cells after axotomy in p75 neurotrophin receptor knockout mice. The unbiased estimation of number and size of arbitrary particles using the disector. A stereological method for estimating the total length and size of myelin fibers in human brain white matter. Methods 73 , — Estimated neuronal populations and volumes of the hippocampus and its subfields in schizophrenia.
Psychiatry , — Stereological principle for morphometry in electron microscopic cytology.