| 1. |
- Woldbye, David P D, et al.
(author)
-
Differential suppression of seizures via Y2 and Y5 neuropeptide Y receptors.
- 2005
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In: Neurobiology of Disease. - : Elsevier BV. - 0969-9961. ; 20:3, s. 760-772
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Journal article (peer-reviewed)abstract
- Neuropepticle Y (NPY) prominently inhibits epileptic seizures in different animal models. The NPY receptors mediating this effect remain controversial partially due to lack of highly selective agonists and antagonists. To circumvent this problem, we used various NPY receptor knockout mice with the same genetic background and explored anti-epileptic action of NPY in vitro and in vivo. In Y2 (Y2-/-) and Y5 (Y5-/-) receptor knockouts, NPY partially inhibited 0 Mg2(+)-induced epileptiform activity in hippocampal slices. In contrast, in double knockouts (Y2Y5-/-), NPY had no effect, suggesting that in the hippocampus in vitro both receptors mediate anti-epileptiform action of NPY in an additive manner. Systemic kainate induced more severe seizures in Y5-/- and Y2Y5-/-, but not in Y2-/- mice, as compared to wild-type mice. Moreover, kainate seizures were aggravated by administration of the Y5 antagonist L-152,804 in wild-type mice. In Y5-/- mice, hippocampal kindling progressed faster, and afterdischarge durations were longer in amygdala, but not in hippocampus, as compared to wild-type controls. Taken together, these data suggest that, in mice, both Y2 and Y5 receptors regulate hippocampa seizures in vitro, while activation of Y5 receptors in extra-hippocampa: regions reduces generalized seizures in vivo.
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| 2. |
- Avaliani, Natalia, et al.
(author)
-
Optogenetics reveal delayed afferent synaptogenesis on grafted human induced pluripotent stem cell-derived neural progenitors.
- 2014
-
In: Stem Cells. - : Oxford University Press (OUP). - 1549-4918 .- 1066-5099. ; 32:12, s. 3088-3098
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Journal article (peer-reviewed)abstract
- Reprogramming of somatic cells into pluripotency stem cell state have opened new opportunities in cell replacement therapy and disease modeling in a number of neurological disorders. It still remains unknown, however, to what degree the grafted human induced pluripotent stem cells (hiPSCs) differentiate into a functional neuronal phenotype and if they integrate into the host circuitry. Here we present a detailed characterization of the functional properties and synaptic integration of hiPSC-derived neurons grafted in an in vitro model of hyperexcitable epileptic tissue, namely organotypic hippocampal slice cultures (OHSC), and in adult rats in vivo. The hiPSCs were first differentiated into long-term self-renewing neuroepithelial stem (lt-NES) cells, which are known to form primarily GABAergic neurons. When differentiated in OHSCs for six weeks, lt-NES cell-derived neurons displayed neuronal properties such as TTX-sensitive sodium currents and action potentials (APs), as well as both spontaneous and evoked postsynaptic currents, indicating functional afferent synaptic inputs. The grafted cells had a distinct electrophysiological profile compared to host cells in the OHSCs with higher input resistance, lower resting membrane potential and APs with lower amplitude and longer duration. To investigate the origin of synaptic afferents to the grafted lt-NES cell-derived neurons, the host neurons were transduced with Channelrhodopsin-2 (ChR2) and optogenetically activated by blue light. Simultaneous recordings of synaptic currents in grafted lt-NES cell-derived neurons using whole-cell patch-clamp technique at 6 weeks after grafting revealed limited synaptic connections from host neurons. Longer differentiation times, up to 24 weeks after grafting in vivo, revealed more mature intrinsic properties and extensive synaptic afferents from host neurons to the It-NES cell-derived neurons, suggesting that these cells require extended time for differentiation/maturation and synaptogenesis. However, even at this later time-point, the grafted cells maintained a higher input resistance. These data indicate that grafted lt-NES cell-derived neurons receive ample afferent input from the host brain. Since the lt-NES cells used in this study show a strong propensity for GABAergic differentiation, the host-to-graft synaptic afferents may facilitate inhibitory neurotransmitter release, and normalize hyperexcitable neuronal networks in brain diseases, e.g. such as epilepsy. Stem Cells 2014.
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| 3. |
- Benson, Thea Emily, et al.
(author)
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Urinary bisphenol a, f and s levels and semen quality in young adult danish men
- 2021
-
In: International Journal of Environmental Research and Public Health. - : MDPI AG. - 1661-7827 .- 1660-4601. ; 18:4
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Journal article (peer-reviewed)abstract
- Bisphenol A (BPA) is considered an endocrine disruptor and has been associated with deleterious effects on spermatogenesis and male fertility. Bisphenol F (BPF) and S (BPS) are struc-turally similar to BPA, but knowledge of their effects on male fertility remains limited. In this cross– sectional study, we investigated the associations between exposure to BPA, BPF, and BPS and semen quality in 556 men 18–20 years of age from the Fetal Programming of Semen Quality (FEPOS) cohort. A urine sample was collected from each participant for determination of BPA, BPF, and BPS concentrations while a semen sample was collected to determine ejaculate volume, sperm concen-tration, total sperm count, sperm motility, and sperm morphology. Associations between urinary bisphenol levels (continuous and quartile–divided) and semen characteristics were estimated using a negative binomial regression model adjusting for urine creatinine concentration, alcohol intake, smoking status, body mass index (BMI), fever, sexual abstinence time, maternal pre–pregnancy BMI, and first trimester smoking, and highest parental education during first trimester. We found no associations between urinary bisphenol of semen quality in a sample of young men from the general Danish population.
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| 4. |
- Berglind, Fredrik, et al.
(author)
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Neuronal activity dynamics in the dentate gyrus during early epileptogenesis
- 2023
-
In: Epilepsy Research. - 0920-1211. ; 194
-
Journal article (peer-reviewed)abstract
- Epileptogenesis is a complex process involving a multitude of changes at the molecular, cellular and network level. Previous studies have identified several key alterations contributing to epileptogenesis and the development of hyper-excitability in different animal models, but only a few have focused on the early stages of this process. For post status epilepticus (SE) temporal lobe epilepsy in particular, understanding network dynamics during the early phases might be crucial for developing accurate preventive treatments to block the development of chronic spontaneous seizures. In this study, we used a viral vector mediated approach to examine activity of neurons in the dentate gyrus of the hippocampus during early epileptogenesis. We find that while granule cells are active 8 h after SE and then gradually decrease their activity, Calretinin-positive mossy cells and Neuropeptide Y-positive GABAergic interneurons in the hilus show a delayed activation pattern starting at 24 and peaking at 48 h after SE. These data suggest that indirect inhibition of granule cells by mossy cells through recruitment of local GABAergic interneurons could be an important mechanisms of excitability control during early epileptogenesis, and contribute to our understanding of the complex role of these cells in normal and pathological conditions.
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| 5. |
- Berglind, Fredrik, et al.
(author)
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Optogenetic inhibition of chemically induced hypersynchronized bursting in mice.
- 2014
-
In: Neurobiology of Disease. - : Elsevier BV. - 0969-9961. ; 65, s. 133-141
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Journal article (peer-reviewed)abstract
- Synchronized activity is common during various physiological operations but can culminate in seizures and consequently in epilepsy in pathological hyperexcitable conditions in the brain. Many types of seizures are not possible to control and impose significant disability for patients with epilepsy. Such intractable epilepsy cases are often associated with degeneration of inhibitory interneurons in the cortical areas resulting in impaired inhibitory drive onto the principal neurons. Recently emerging optogenetic technique has been proposed as an alternative approach to control such seizures but whether it may be effective in situations where inhibitory processes in the brain are compromised has not been addressed. Here we used pharmacological and optogenetic techniques to block inhibitory neurotransmission and induce epileptiform activity in vitro and in vivo. We demonstrate that NpHR-based optogenetic hyperpolarization and thereby inactivation of a principal neuronal population in the hippocampus is effectively attenuating seizure activity caused by disconnected network inhibition both in vitro and in vivo. Our data suggest that epileptiform activity in the hippocampus caused by impaired inhibition may be controlled by optogenetic silencing of principal neurons and potentially can be developed as an alternative treatment for epilepsy.
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| 6. |
- Gaml-Sørensen, Anne, et al.
(author)
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Maternal vitamin D levels and male reproductive health : a population-based follow-up study
- 2023
-
In: European Journal of Epidemiology. - : Springer Science and Business Media LLC. - 0393-2990 .- 1573-7284. ; 38:5, s. 469-484
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Journal article (peer-reviewed)abstract
- Maternal vitamin D levels during pregnancy may be important for reproductive health in male offspring by regulating cell proliferation and differentiation during development. We conducted a follow-up study of 827 young men from the Fetal Programming of Semen Quality (FEPOS) cohort, nested in the Danish National Birth Cohort to investigate if maternal vitamin D levels were associated with measures of reproductive health in adult sons. These included semen characteristics, testes volume, and reproductive hormone levels and were analysed according to maternal vitamin D (25(OH)D3) levels during pregnancy. In addition, an instrumental variable analysis using seasonality in sun exposure as an instrument for maternal vitamin D levels was conducted. We found that sons of mothers with vitamin D levels < 25 nmol/L had 11% (95% CI − 19 to − 2) lower testes volume and a 1.4 (95% CI 1.0 to 1.9) times higher risk of having low testes volume (< 15 mL), in addition to 20% (95% CI − 40 to 9) lower total sperm count and a 1.6 (95% CI 0.9 to 2.9) times higher risk of having a low total sperm count (< 39 million) compared with sons of mothers with vitamin D levels > 75 nmol/L. Continuous models, spline plots and an instrumental variable analysis supported these findings. Low maternal vitamin D levels were associated with lower testes volume and lower total sperm count with indications of dose-dependency. Maternal vitamin D level above 75 nmol/L during pregnancy may be beneficial for testes function in adult sons.
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| 7. |
- Gaml-Sørensen, Anne, et al.
(author)
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The estimated effect of season and vitamin D in the first trimester on pubertal timing in girls and boys : A cohort study and an instrumental variable analysis
- 2023
-
In: International Journal of Epidemiology. - 0300-5771. ; 52:5, s. 1328-1340
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Journal article (peer-reviewed)abstract
- Background: Season of birth has been associated with age at menarche. Maternal vitamin D levels in pregnancy may explain this effect. We investigated whether the season of first trimester or maternal 25-hydroxyvitamin D3 [25(OH)D3] levels were associated with pubertal timing in children. Methods: We conducted a follow-up study of 15 819 children born in 2000-03 from the Puberty Cohort, nested in the Danish National Birth Cohort (DNBC). Mean differences in attaining numerous pubertal markers, including a combined estimate for the average age at attaining all pubertal markers, were estimated for low (November-April) relative to high (May-October) sunshine exposure season in the first trimester using multivariable interval-censored regression models. Moreover, we conducted a two-sample instrumental variable analysis using season as an instrument for maternal first-Trimester 25(OH)D3 plasma levels obtained from a non-overlapping subset (n = 827) in the DNBC. Results: For the combined estimate, girls and boys of mothers who had their first trimester during November-April had earlier pubertal timing than girls and boys of mothers whose first trimester occurred during May-October:-1.0 months (95% CI:-1.7 to-0.3) and-0.7 months (95% CI:-1.4 to-0.1), respectively. In the instrumental variable analysis, girls and boys also had earlier pubertal timing: respectively,-1.3 months (95% CI:-2.1 to-0.4) and-1.0 months (95% CI:-1.8 to-0.2) per SD (22 nmol/L) decrease in 25(OH)D3. Conclusions: Both first pregnancy trimester during November-April and lower 25(OH)D3 were associated with earlier pubertal timing in girls and boys.
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| 8. |
- Gotzsche, Casper R., et al.
(author)
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Combined gene overexpression of neuropeptide Y and its receptor Y5 in the hippocampus suppresses seizures
- 2012
-
In: Neurobiology of Disease. - : Elsevier BV. - 0969-9961. ; 45:1, s. 288-296
-
Journal article (peer-reviewed)abstract
- We recently demonstrated that recombinant adeno-associated viral vector-induced hippocampal overexpression of neuropeptide Y receptor, Y2, exerts a seizure-suppressant effect in kindling and kainate-induced models of epilepsy in rats. Interestingly, additional overexpression of neuropeptide Y in the hippocampus strengthened the seizure-suppressant effect of transgene Y2 receptors. Here we show for the first time that another neuropeptide Y receptor, Y5, can also be overexpressed in the hippocampus. However, unlike Y2 receptor overexpression, transgene Y5 receptors in the hippocampus had no effect on kainate-induced motor seizures in rats. However, combined overexpression of Y5 receptors and neuropeptide Y exerted prominent suppression of seizures. This seizure-suppressant effect of combination gene therapy with Y5 receptors and neuropeptide Y was significantly stronger as compared to neuropeptide Y overexpression alone. These results suggest that overexpression of Y5 receptors in combination with neuropeptide Y could be an alternative approach for more effective suppression of hippocampal seizures. (C) 2011 Elsevier Inc. All rights reserved.
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| 9. |
- Kanter Schlifke, Irene, et al.
(author)
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Galanin gene transfer curtails generalized seizures in kindled rats without altering hippocampal synaptic plasticity
- 2007
-
In: Neuroscience. - : Elsevier BV. - 1873-7544 .- 0306-4522. ; 150:4, s. 984-992
-
Journal article (peer-reviewed)abstract
- Gene therapy-based overexpression of endogenous seizure-suppressing molecules represents a promising treatment strategy for epilepsy. Viral vector-based overexpression of the neuropeptide galanin has been shown to effectively suppress generalized seizures in various animal models of epilepsy. However, it has not been explored whether such treatment can also prevent the epileptogenesis. Using a recombinant adeno-associated viral (rAAV) vector, we induced hippocampal galanin overexpression under the neuron specific enolase promoter in rats. Here we report that in animals with galanin overexpression, the duration of electrographic afterdischarges was shortened and initiation of convulsions was delayed at generalized seizure stages. However, the hippocampal kindling development was unchanged. Short-term plasticity of mossy fiber-cornu ammonis (CA) 3 synapses was unaltered, as assessed by paired-pulse and frequency facilitation of field excitatory postsynaptic potentials (fEPSPs) in hippocampal slices, suggesting that despite high transgene galanin expression, overall release probability of glutamate in these synapses was unaffected. These data indicate that hippocampal rAAV-based galanin overexpression is capable of mediating anticonvulsant effects by lowering the seizure susceptibility once generalized seizures are induced, but does not seem to affect kindling development or presynaptic short-term plasticity in mossy fibers.
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| 10. |
- Kokaia, Merab, et al.
(author)
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Epilepsy and Gene Therapy: Resculpturing Synaptic Transmission with Neuropeptides
- 2009
-
In: Encyclopedia of Basic Epilepsy Research. ; , s. 1430-1434
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Book chapter (peer-reviewed)abstract
- One of the most promising and novel strategies to interfere with neurological disease processes is gene therapy using recombinant adeno-associated viral (rAAV) vectors. Such a rAAV-based gene delivery approach is rapidly advancing towards clinical trials. In this regard, neuropeptide Y (NPY) gene transduction into the brain tissue has attracted particular interest due to its potential to regulate and perhaps even ameliorate epileptic conditions. NPY gene transduction by viral vectors in epileptogenic regions of the brain can effectively suppress seizures in animals. The mechanisms underlying the seizure-suppressant effects of an NPY transgene are not well understood; in particular, under which circumstances transgene NPY is released, and whether and how it acts on synaptic transmission within the area of viral vector transduction are not known. These questions are of fundamental importance not only for the implementation of such a gene therapy approach in clinical trials with patients, but also for our general understanding of how transgene neuropeptides may act in the brain.
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| 11. |
- Kokaia, Merab, et al.
(author)
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The treatment of neurological diseases under a new light: the importance of optogenetics.
- 2011
-
In: Drugs of Today. - : Clarivate Analytics (US). - 1699-3993. ; 47:1, s. 53-62
-
Research review (peer-reviewed)abstract
- Controlling activity of defined populations of neurons without affecting other neurons in the brain is now possible by a new gene- and neuroengineering technology termed optogenetics. Derived from microbial organisms, opsin genes encoding light-activated ion channels and pumps (channelrhodopsin-2 [ChR2]; halorhodopsin [NpHR], respectively), engineered for expression in the mammalian brain, can be genetically targeted into specific neural populations using viral vectors. When exposed to light with appropriate wavelength, action potentials can be triggered in ChR2-expressing neurons, whereas inhibition of action potentials can be obtained in NpHR-expressing neurons, thus allowing for powerful control of neural activity. Optogenetics is now intensively used in laboratory animals, both in vitro and in vivo, for exploring functions of complex neural circuits and information processing in the normal brain and during various neurological conditions. The clinical perspectives of adopting optogenetics as a novel treatment strategy for human neurological disorders have generated considerable interest, largely because of the enormous potential demonstrated in recent rodent and nonhuman primate studies. Restoration of dopamine-related movement dysfunction in parkinsonian animals, amelioration of blindness and recovery of breathing after spinal cord injury are a few examples of such perspectives.
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| 12. |
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| 13. |
- Ledri, Marco, et al.
(author)
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Tuning afferent synapses of hippocampal interneurons by neuropeptide Y.
- 2011
-
In: Hippocampus. - : Wiley. - 1050-9631. ; 21, s. 198-211
-
Journal article (peer-reviewed)abstract
- Cholecystokinin (CCK)-expressing basket cells encompass a subclass of inhibitory GABAergic interneurons that regulate memory-forming oscillatory network activity of the hippocampal formation in accordance to the emotional and motivational state of the animal, conveyed onto these cells by respective extrahippocampal afferents. Various excitatory and inhibitory afferent and efferent synapses of the hippocampal CCK basket cells express serotoninergic, cholinergic, cannabinoid, and benzodiazepine sensitive receptors, all contributing to their functional plasticity. We explored whether CCK basket cells are modulated by neuropeptide Y (NPY), one of the major local neuropeptides that strongly inhibits hippocampal excitability and has significant effect on its memory function. Here, using GAD65-GFP transgenic mice for prospective identification of CCK basket cells and whole-cell patch-clamp recordings, we show for the first time that excitatory and inhibitory inputs onto CCK basket cells in the dentate gyrus of the hippocampus are modulated by NPY through activation of NPY Y2 receptors. The frequency of spontaneous and miniature EPSCs, as well as the amplitudes of stimulation-evoked EPSCs were decreased. Similarly, the frequency of both spontaneous and miniature IPSCs, and the amplitudes of stimulation-evoked IPSCs were decreased after NPY application. Most of the effects of NPY could be attributed to a presynaptic site of action. Our data provide the first evidence that the excitatory and inhibitory inputs onto the CCK basket cells could be modulated by local levels of NPY, and may change the way these cells process extrahippocampal afferent information, influencing hippocampal function and its network excitability during normal and pathological oscillatory activities. (c) 2009 Wiley-Liss, Inc.
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| 14. |
- Noe, Francesco M., et al.
(author)
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Gene therapy of focal-onset epilepsy by adeno-associated virus vector-mediated overexpression of neuropeptide Y
- 2010
-
In: Epilepsia. - : Wiley. - 0013-9580. ; 51, s. 96-96
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Journal article (peer-reviewed)abstract
- P>Adeno-associated virus (AAV)-mediated neuropeptide Y (NPY) overexpression in areas of seizure onset or generalization may be effective for the treatment of pharmacoresistant seizures in focal onset epilepsy. NPY overexpression mediates anticonvulsant activity in various seizures models and antiepileptogenic effects in kindling. Side effects are limited thus suggesting that this therapeutic approach could be effective and relatively safe. For an expanded treatment of this topic see Jasper's basic mechanisms of the epilepsies. 4th ed. (Noebels JL, Avoli M, Rogawski MA, Olsen RW, Delgado-Escueta AV, eds) published by Oxford University Press (available on the National Library of Medicine Bookshelf [NCBI] at http://www.ncbi.nlm.nih.gov/books).
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| 15. |
- Parish, CL, et al.
(author)
-
Wnt5a-treated midbrain neural stem cells improve dopamine cell replacement therapy in parkinsonian mice
- 2008
-
In: Journal of Clinical Investigation. - 0021-9738. ; 118:1, s. 149-160
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Journal article (peer-reviewed)abstract
- Dopamine (DA) cell replacement therapy in Parkinson disease (PD) can be achieved using human fetal mesencephalic tissue; however, limited tissue availability has hindered further developments. Embryonic stem cells provide a promising alternative, but poor survival and risk of teratoma formation have prevented their clinical application. We present here a method for generating large numbers of DA neurons based on expanding and differentiating ventral midbrain (VM) neural stem cells/progenitors in the presence of key signals necessary for VM DA neuron development. Mouse VM neurospheres (VMNs) expanded with FGF2, differentiated with sonic hedgehog and FGF8, and transfected with Wnt5a (VMN-Wnt5a) generated 10-fold more DA neurons than did conventional FGF2-treated VMNs. VMN-Wnt5a cells exhibited the transcriptional and biochemical profiles and intrinsic electrophysiological properties of midbrain DA cells. Transplantation of these cells into parkinsonian mice resulted in significant cellular and functional recovery. Importantly, no tumors were detected and only a few transplanted grafts contained sporadic nestin-expressing progenitors. Our findings show that Wnt5a improves the differentiation and functional integration of stem cell-derived DA neurons in vivo and define Wnt5a-treated neural stem cells as an efficient and safe source of DA neurons for cell replacement therapy in PD.
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| 16. |
- Toft Sörensen, Andreas, et al.
(author)
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Activity-dependent long-term plasticity of afferent synapses on grafted stem/progenitor cell-derived neurons.
- 2011
-
In: Experimental Neurology. - : Elsevier BV. - 0014-4886. ; 229, s. 274-281
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Journal article (peer-reviewed)abstract
- Stem cell-based cell replacement therapies aiming at restoring injured or diseased brain function ultimately rely on the capability of transplanted cells to promote functional recovery. The mechanisms by which stem cell-based therapies for neurological conditions can lead to functional recovery are uncertain, but structural and functional repair appears to depend on integration of transplanted cell-derived neurons into neuronal circuitries. The nature by which stem/progenitor cell-derived neurons synaptically integrate into neuronal circuitries is largely unexplored. Here we show that transplanted GFP-labeled neuronal progenitor cells into the rat hippocampus exhibit mature neuronal morphology following 4-10 weeks. GFP-positive cells were preferentially integrated into the principal cell layers of hippocampus, particularly CA3. Patch-clamp recordings from GFP-expressing cells revealed that they generated fast action potentials, and their intrinsic membrane properties were overall similar to endogenous host neurons recorded in same areas. As judged by occurrence of spontaneous excitatory postsynaptic currents (EPSCs), transplanted GFP-positive cells were synaptically integrated into the host circuitry. Comparable to host neurons, both paired-pulse depression and facilitation of afferent fiber stimulation-evoked EPSCs were observed in GFP-positive cells. Upon high-frequency stimulation, GFP-positive cells displayed post-tetanic potentiation of EPSCs, in some cases followed by long-term potentiation (LTP) lasting for more than 30 min. Our data show for the first time that transplanted neuronal progenitor cells can become functional neurons and their afferent synapses are capable of expressing activity-dependent short and long-term plasticity. These synaptic properties may facilitate host-to-graft interactions and regulate activity of the grafted cells promoting functional recovery of the diseased brain.
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| 17. |
- Toft Sörensen, Andreas, et al.
(author)
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Activity-dependent volume transmission by transgene NPY attenuates glutamate release and LTP in subiculum
- 2008
-
In: Molecular and Cellular Neuroscience. - : Elsevier BV. - 1044-7431. ; 39:2, s. 37-229
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Journal article (peer-reviewed)abstract
- Neuropeptide Y (NPY) gene transduction of the brain using viral vectors in epileptogenic regions can effectively suppress seizures in animals, and is being considered as a promising alternative treatment strategy for epilepsy. Therefore, it is fundamental to understand the detailed mechanisms governing the release and action of transgene NPY in neuronal circuitries. Using whole-cell recordings from subicular neurons, we show that in animals transduced by recombinant adeno-associated viral (rAAV) vector carrying the NPY gene, transgene NPY is released during high-frequency activation of CA1-subicular synapses. Released transgene NPY attenuates excitatory synaptic transmission not only in activated, but also in neighboring, non-activated synapses. Such broad action of transgene NPY may prevent recruitment of excitatory synapses in epileptic activity and could play a key role in limiting the spread and generalization of seizures.
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| 18. |
- Toft Sörensen, Andreas, et al.
(author)
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Functional properties and synaptic integration of genetically labelled dopaminergic neurons in intrastriatal grafts.
- 2005
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In: European Journal of Neuroscience. - : Wiley. - 1460-9568 .- 0953-816X. ; 21:10, s. 2793-2799
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Journal article (peer-reviewed)abstract
- ntrastriatal grafts of fetal ventral mesencephalic tissue, rich in dopaminergic neurons, can reverse symptoms in Parkinson's disease. For development of effective cell replacement therapy, other sources of dopaminergic neurons, e.g. derived from stem cells, are needed. However, the electrophysiological properties grafted cells need to have in order to induce substantial functional recovery are poorly defined. It has not been possible to prospectively identify and record from dopaminergic neurons in fetal transplants. Here we used transgenic mice expressing green fluorescent protein under control of the rat tyrosine hydroxylase promoter for whole-cell patch-clamp recordings of endogenous and grafted dopaminergic neurons. We transplanted ventral mesencephalic tissue from E12.5 transgenic mice into striatum of neonatal rats with or without lesions of the nigrostriatal dopamine system. The transplanted cells exhibited intrinsic electrophysiological properties typical of substantia nigra dopaminergic neurons, i.e. broad action potentials, inward rectifying currents with characteristic 'sag', and spontaneous action potentials. The grafted dopaminergic neurons also received functional excitatory and inhibitory synaptic inputs from the host brain, as shown by the presence of both spontaneous and stimulation-evoked excitatory and inhibitory postsynaptic currents. Occurrence of spontaneous excitatory and inhibitory currents was lower, and of spontaneous action potentials was higher, in neurons placed in the dopamine-depleted striatum than of those in the intact striatum. Our findings define specific electrophysiological characteristics of transplanted fetal dopaminergic neurons, and we provide the first direct evidence of functional synaptic integration of these neurons into host neural circuitries.
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| 19. |
- Toft Sörensen, Andreas
(author)
-
Gene therapy for epilepsy: resculpturing synaptic transmission with neuropeptides
- 2008
-
Doctoral thesis (other academic/artistic)abstract
- Intractable seizures and lack of effective antiepileptic drugs (AED) are severe and common conditions affecting many patients with epilepsy. Thus, there is an urgent need to develop new therapies in epilepsy. The search for novel treatments has identified several neuropeptide systems as potential targets for future therapeutic interventions. The neuropeptide Y (NPY) may represent one such target, as it plays a key role in controlling excitability in the hippocampus by suppressing glutamatergic transmission. If manipulated, the NPY system may be capable of restoring the imbalance between excitation and inhibition occurring in the epileptic brain. Indeed, emerging evidence has established a proof-of-principle for viral vector-mediated transfer of gene and expression of NPY in epileptogenic regions of the brain, providing effective suppression of both acute and chronic seizures in animal models of epilepsy. Therefore, NPY gene therapy strategies are currently under intense investigation and clinical trials are forthcoming. To implement an effective NPY gene therapy in patients, as well as to extend our general knowledge of how transgene NPY may act in the brain, the receptor subtypes mediating the antiepileptic action of NPY needs to be determined. Moreover, the mechanisms underlying the seizure-suppressant effects of transgene NPY are not well understood. Particularly, we need to know under which circumstances transgene NPY is released, and whether and how it acts on synaptic transmission within the area of viral vector transduction. In this thesis, evidences are provided showing that NPY is mediating its antiepileptic effect through activation of both Y2 and Y5 receptors in the hippocampus, and predominately via Y5 receptors in extra-hippocampal areas. Moreover, in rats, hippocampal NPY gene therapy generates long-lasting and neuronal-specific overexpression of transgene NPY. This is not associated with alterations in basal synaptic transmission, probably due to minor constitutive release of transgene NPY. However, as transgene NPY is preferentially released during high frequency neuronal activity, acting as a volume transmitter, it interferes with neuronal activity-dependent processes, reflected by suppressed long-lasting synaptic plasticity in hippocampal synapses and delayed, but not impaired, hippocampal dependent learning capacity in naïve animals. This could be an unwanted adverse effect of transgene NPY, but since already existing impairment of long-term synaptic plasticity is not further exacerbated after electrical kindling-induced seizures, NPY gene therapy still remains a promising and novel antiepileptic treatment strategy.
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| 20. |
- Toft Sörensen, Andreas, et al.
(author)
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Hippocampal NPY gene transfer attenuates seizures without affecting epilepsy-induced impairment of LTP.
- 2009
-
In: Experimental Neurology. - : Elsevier BV. - 0014-4886. ; 215, s. 328-333
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Journal article (peer-reviewed)abstract
- Recently, hippocampal neuropeptide Y (NPY) gene therapy has been shown to effectively suppress both acute and chronic seizures in animal model of epilepsy, thus representing a promising novel antiepileptic treatment strategy, particularly for patients with intractable mesial temporal lobe epilepsy (TLE). However, our previous studies show that recombinant adeno-associated viral (rAAV)-NPY treatment in naive rats attenuates long-term potentiation (LTP) and transiently impairs hippocampal learning process, indicating that negative effect on memory function could be a potential side effect of NPY gene therapy. Here we report how rAAV vector-mediated overexpression of NPY in the hippocampus affects rapid kindling, and subsequently explore how synaptic plasticity and transmission is affected by kindling and NPY overexpression by field recordings in CA1 stratum radiatum of brain slices. In animals injected with rAAV-NPY, we show that rapid kindling-induced hippocampal seizures in vivo are effectively suppressed as compared to rAAV-empty injected (control) rats. Six to nine weeks later, basal synaptic transmission and short-term synaptic plasticity are unchanged after rapid kindling, while LTP is significantly attenuated in vitro. Importantly, transgene NPY overexpression has no effect on short-term synaptic plasticity, and does not further compromise LTP in kindled animals. These data suggest that epileptic seizure-induced impairment of memory function in the hippocampus may not be further affected by rAAV-NPY treatment, and may be considered less critical for clinical application in epilepsy patients already experiencing memory disturbances.
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| 21. |
- Toft Sörensen, Andreas, et al.
(author)
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NPY gene transfer in the hippocampus attenuates synaptic plasticity and learning.
- 2008
-
In: Hippocampus. - : Wiley. - 1050-9631 .- 1098-1063. ; 18:6, s. 564-574
-
Journal article (peer-reviewed)abstract
- Recombinant adeno-associated viral (rAAV) vector-induced neuropeptide Y (NPY) overexpression in the hippocampus exerts powerful antiepileptic and antiepileptogenic effects in rats. Such gene therapy approach could be a valuable alternative for developing new antiepileptic treatment strategies. Future clinical progress, however, requires more detailed evaluation of possible side effects of this treatment. Until now it has been unknown whether rAAV vector-based NPY overexpression in the hippocampus alters normal synaptic transmission and plasticity, which could disturb learning and memory processing. Here we show, by electrophysiological recordings in CA1 of the hippocampal formation of rats, that hippocampal NPY gene transfer into the intact brain does not affect basal synaptic transmission, but slightly alters short-term synaptic plasticity, most likely via NPY Y2 receptor-mediated mechanisms. In addition, transgene NPY seems to be released during high frequency neuronal activity, leading to decreased glutamate release in excitatory synapses. Importantly, memory consolidation appears to be affected by the treatment. We found that long-term potentiation (LTP) in the CA1 area is partially impaired and animals have a slower rate of hippocampal-based spatial discrimination learning. These data provide the first evidence that rAAV-based gene therapy using NPY exerts relative limited effect on synaptic plasticity and learning in the hippocampus, and therefore this approach could be considered as a viable alternative for epilepsy treatment. (c) 2008 Wiley-Liss, Inc.
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| 22. |
- Tønnesen, Jan, et al.
(author)
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Functional integration of grafted neural stem cell-derived dopaminergic neurons monitored by optogenetics in an in vitro Parkinson model.
- 2011
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In: PLoS ONE. - : Public Library of Science (PLoS). - 1932-6203. ; 6:3
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Journal article (peer-reviewed)abstract
- Intrastriatal grafts of stem cell-derived dopamine (DA) neurons induce behavioral recovery in animal models of Parkinson's disease (PD), but how they functionally integrate in host neural circuitries is poorly understood. Here, Wnt5a-overexpressing neural stem cells derived from embryonic ventral mesencephalon of tyrosine hydroxylase-GFP transgenic mice were expanded as neurospheres and transplanted into organotypic cultures of wild type mouse striatum. Differentiated GFP-labeled DA neurons in the grafts exhibited mature neuronal properties, including spontaneous firing of action potentials, presence of post-synaptic currents, and functional expression of DA D(2) autoreceptors. These properties resembled those recorded from identical cells in acute slices of intrastriatal grafts in the 6-hydroxy-DA-induced mouse PD model and from DA neurons in intact substantia nigra. Optogenetic activation or inhibition of grafted cells and host neurons using channelrhodopsin-2 (ChR2) and halorhodopsin (NpHR), respectively, revealed complex, bi-directional synaptic interactions between grafted cells and host neurons and extensive synaptic connectivity within the graft. Our data demonstrate for the first time using optogenetics that ectopically grafted stem cell-derived DA neurons become functionally integrated in the DA-denervated striatum. Further optogenetic dissection of the synaptic wiring between grafted and host neurons will be crucial to clarify the cellular and synaptic mechanisms underlying behavioral recovery as well as adverse effects following stem cell-based DA cell replacement strategies in PD.
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| 23. |
- Tønnesen, Jan, et al.
(author)
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Optogenetic control of epileptiform activity.
- 2009
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In: Proceedings of the National Academy of Sciences. - : Proceedings of the National Academy of Sciences. - 1091-6490 .- 0027-8424. ; 106:29, s. 12162-12167
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Journal article (peer-reviewed)abstract
- The optogenetic approach to gain control over neuronal excitability both in vitro and in vivo has emerged as a fascinating scientific tool to explore neuronal networks, but it also opens possibilities for developing novel treatment strategies for neurologic conditions. We have explored whether such an optogenetic approach using the light-driven halorhodopsin chloride pump from Natronomonas pharaonis (NpHR), modified for mammalian CNS expression to hyperpolarize central neurons, may inhibit excessive hyperexcitability and epileptiform activity. We show that a lentiviral vector containing the NpHR gene under the calcium/calmodulin-dependent protein kinase IIalpha promoter transduces principal cells of the hippocampus and cortex and hyperpolarizes these cells, preventing generation of action potentials and epileptiform activity during optical stimulation. This study proves a principle, that selective hyperpolarization of principal cortical neurons by NpHR is sufficient to curtail paroxysmal activity in transduced neurons and can inhibit stimulation train-induced bursting in hippocampal organotypic slice cultures, which represents a model tissue of pharmacoresistant epilepsy. This study demonstrates that the optogenetic approach may prove useful for controlling epileptiform activity and opens a future perspective to develop it into a strategy to treat epilepsy.
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| 24. |
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| 25. |
- Woldbye, David P D, et al.
(author)
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Adeno-associated viral vector-induced overexpression of neuropeptide Y Y2 receptors in the hippocampus suppresses seizures.
- 2010
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In: Brain. - : Oxford University Press (OUP). - 1460-2156 .- 0006-8950. ; 133:Pt 9, s. 2778-2788
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Journal article (peer-reviewed)abstract
- Gene therapy using recombinant adeno-associated viral vectors overexpressing neuropeptide Y in the hippocampus exerts seizure-suppressant effects in rodent epilepsy models and is currently considered for clinical application in patients with intractable mesial temporal lobe epilepsy. Seizure suppression by neuropeptide Y in the hippocampus is predominantly mediated by Y2 receptors, which, together with neuropeptide Y, are upregulated after seizures as a compensatory mechanism. To explore whether such upregulation could prevent seizures, we overexpressed Y2 receptors in the hippocampus using recombinant adeno-associated viral vectors. In two temporal lobe epilepsy models, electrical kindling and kainate-induced seizures, vector-based transduction of Y2 receptor complementary DNA in the hippocampus of adult rats exerted seizure-suppressant effects. Simultaneous overexpression of Y2 and neuropeptide Y had a more pronounced seizure-suppressant effect. These results demonstrate that overexpression of Y2 receptors (alone or in combination with neuropeptide Y) could be an alternative strategy for epilepsy treatment.
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