Neue Ergebnisse zur morphologischen Charakterisierung der enterischen Coinnervation der quergestreiften Ösophagusmuskulatur.

Background and Aim In the present work, we focused on enteric coinnervation of striated muscle fibers in the esophagus. Enteric coinnervation has been demonstrated in several species including human. For example, the esophagus of phylogenetically old mammals like the bat and also ruminants like the antelope and he-goat show this innervation scheme. This work was undertaken to investigate esophagi of sheep and shrew to complete the category of ruminants and old mammals. As little is known about the morphological organization of serotoninergic neurons and other serotonin-containing elements in the esophagus, the second aim of this study was to characterize the serotonin (5-HT) system in the mouse esophagus. Several neuronal markers, e.g. vasoactive intestinal peptide (VIP), choline acetyltransferase (ChAT) and tyrosine hydroxylase (TH) were tested to clarify colocalization with serotonin. In addition, the issue of intrinsic versus extrinsic origin of 5-HT was investigated. Methods Experiments were performed on 76 adult and three juvenile mice (C57/Bl6), one sheep and three shrews. Furthermore, 12 tryptophan hydroxylase (TPH)-knockout-mice were included. The tissues of some mice was excised, incubated in oxygenated Krebs-Ringer solution, and subsequently fixed in 4 % phosphate-buffered formaldehyde. Other mice were perfused with 4 % phosphate-buffered formaldehyde before organ excision without preincubation in Krebs-Ringer solution. Cryosections were performed of esophagi, small intestine and brain. Whole-mounts were prepared of esophagi and small intestine. All samples were investigated histologically and by enzyme- and immunohistochemistry. Results We could demonstrate enteric coinnervation of striated muscle fibers in the esophagus of sheep and shrew. 5-HT-positive nerve fibers were present in the lamina muscularis mucosae, tunica muscularis, in the myenteric plexus and around blood vessels. Mast-cells in the tela submucosa und tunica adventitia were also positive for 5-HT. On average, about 10 % of all myenteric neurons were stained for 5-HT and about 13 % of all motor endplates were co-innervated by enteric nerve terminals positive for 5-HT. We found about 30 % 5-HT/VIP cell bodies, 3 % 5-HT/ChAT and 6 % 5-HT/TH neurons in the myenteric plexus. On motor endplates, we detected about 5 % 5-HT/VIP- and 64 % 5-HT/TH-positive nerve fibers. Nerve cell bodies of the nucleus ambiguus were negative for 5-HT, but were endowed with 5-HT-positive nerve fibers. These fibers probably derive from 5-HT-positive neurons of the raphe nuclei. The question, whether serotonin is also produced in enteric neurons could not definitely be resolved. Antibodies against TPH did not show adequate staining and even cross reactivity with TH. Investigation of TPH-knockout-mice provided ambiguous results. Conclusion This study provides further information about the occurrence and potential function of enteric coinnervation of esophageal striated muscle fibers. We identified enteric coinnervation in the esophagus of additional species and established serotonin as a new component in this innervation pattern. Serotonin might modulate vagal innervation of esophageal striated muscles both at the central level via projections of raphe nuclei to the compact formation of the nucleus ambiguus and at the peripheral level via enteric coinnervation of motor endplates. The characterization of serotonin in the esophagus helps to interpret previous pharmacological and functional data. However, we were not able to definitely clarify the question, whether serotonin is a genuine product of myenteric neurons. At least some 5-HT-positive nerve fibers in the esophagus may be in fact of sympathetic origin.Hintergrund und Ziele Ein Schwerpunkt der Arbeit stellt die weitere Untersuchung der enterischer Coinnervation an quergestreifter Muskulatur der Speiseröhre dar. Dieses spezielle Innervationsschema wurde an verschiedenen Tierarten sowie beim Menschen nachgewiesen. So zeigen phylogenetisch alte Säuger wie die Fledermaus, aber auch Wiederkäuer wie Antilope und Schraubenziege, enterische Coinnervation. In dieser Arbeit vervollständigten wir die Tiergruppen, indem zusätzlich Speiseröhren von Spitzmaus und Schaf untersucht und quantifiziert wurden. Da es im Gegensatz zu pharmakologischen und funktionellen Untersuchungen kaum morphologische Daten zu Serotonin (5-HT) in der Speiseröhre gibt, fokussiert diese Arbeit vor allem auch auf dem Nachweis von Serotonin in der Speiseröhre der Maus. Antikörper gegen verschiedene neuronale Marker (vasoaktives intestinales Peptid (VIP), Cholinazetyltransferase (ChAT), Tyrosinhydroxylase (TH)) wurden benutzt, um festzustellen, mit welchen Markern Serotonin möglicherweise colokalisiert ist. Weiterhin galt es zu klären, ob Serotonin tatsächlich in enterischen Neuronen produziert oder aus externen Quellen aufgenommen wird. Methoden Die Untersuchungen wurden an 76 adulten und drei juvenilen C57/Bl6 Mäusen, einem Schaf und drei Spitzmäusen durchgeführt. Außerdem wurde Gewebe von 12 Tryptophanhydroxylase (TPH)-Knockout-Mäusen untersucht. Bei einem Teil der Mäuse wurde das Material unfixiert entnommen, in begaster Krebs-Ringer-Lösung inkubiert und anschließend mit 4 % Paraformaldehyd (PFA) immersionsfixiert. Ein Teil der Mäuse wurde mit 4 % PFA perfusionsfixiert und anschließend das Gewebe entnommen. An Gefrierschnitten von Speiseröhre, Dünndarm und Gehirn sowie an Häutchenpräparaten von Speiseröhre und Dünndarm wurden sowohl histologische als auch enzym- bzw. immunhistochemische Färbungen durchgeführt, ausgewertet und dokumentiert. Ergebnisse Enterische Coinnervation der quergestreiften Muskulatur der Speiseröhre konnte bei Spitzmaus und Schaf nachgewiesen werden. 5-HT-positive Nervenfasern konnten in der Lamina muscularis mucosae, der Tunica muscularis, im myenterischen Plexus, sowie um Blutgefäße nachgewiesen werden. Des Weiteren konnten wir stark 5-HT-immunoreaktive Mastzellen in der Tela submucosa und Tunica adventitia darstellen. Etwa 10 % der enterischen Neurone waren 5-HT-positiv. Etwa 13 % aller motorischen Endplatten wurden von 5-HT-positiven Nervenendigungen kontaktiert. Die Anzahl der 5-HT-positiven Neurone, die mit VIP colokalisiert waren lag bei ca. 30 %, mit ChAT zeigten lediglich 3 % und mit TH 6 % der 5-HT-positiven Neurone Colokalisation. An motorischen Endplatten waren ca. 5 % aller 5-HT-positiven Nervenfasern auch VIP-immunreaktiv sowie 64 % TH-positiv. Im Nucleus ambiguus waren keine Nervenzellkörper positiv für 5-HT, jedoch zahlreiche Nervenfasern, welche vermutlich aus den 5-HT-positiven Neuronen der Raphekerne stammen. Ob Serotonin in enterischen Neuronen der Speiseröhre produziert wird, konnte nicht eindeutig nachgewiesen werden. Antikörper gegen TPH zeigten keine zufriedenstellende Färbung bzw. reagierten kreuz mit TH. Selbst Untersuchungen an TPH-Knockout-Mäuse lieferten kein eindeutiges Ergebnis. Schlussfolgerung Kenntnisse über das Auftreten enterischer Coinnervation bei unterschiedlichen Spezies und Serotonin als neuen und weiteren Transmitter dieses Innervationsschemas liefern uns weitere wertvolle Informationen über das funktionelle Ausmaß enterischer Modulation der quergestreiften Ösophagusmuskulatur. Serotonin dürfte die quergestreifte Speiseröhrenmuskulatur nicht nur auf zentraler Ebene via Projektionen der Raphekerne zum Nucleus ambiguus, sondern auch peripher via enterischer Coinnervation beeinflussen. Weiterhin liefern wir mit der Charakterisierung von Serotonin in der Speiseröhre der Maus neue Aspekte, welche bisherige pharmakologische und funktionelle Daten besser interpretieren helfen. Ob Serotonin in der Speiseröhre tatsächlich intrinsischen Ursprungs ist, konnte nicht abschließend geklärt werden. Zumindest ein Teil der 5-HT-positiven Nervenfasern in der Speiseröhre dürfte jedoch sympathischer Herkunft sein

First Study of Combined Blazar Light Curves with FACT and HAWC

For studying variable sources like blazars, it is crucial to achieve unbiased monitoring, either with dedicated telescopes in pointing mode or survey instruments. At TeV energies, the High Altitude Water Cherenkov (HAWC) observatory monitors approximately two thirds of the sky every day. It uses the water Cherenkov technique, which provides an excellent duty cycle independent of weather and season. The First G-APD Cherenkov Telescope (FACT) monitors a small sample of sources with better sensitivity, using the imaging air Cherenkov technique. Thanks to its camera with silicon-based photosensors, FACT features an excellent detector performance and stability and extends its observations to times with strong moonlight, increasing the duty cycle compared to other imaging air Cherenkov telescopes. As FACT and HAWC have overlapping energy ranges, a joint study can exploit the longer daily coverage given that the observatories' locations are offset by 5.3 hours. Furthermore, the better sensitivity of FACT adds a finer resolution of features on hour-long time scales, while the continuous duty cycle of HAWC ensures evenly sampled long-term coverage. Thus, the two instruments complement each other to provide a more complete picture of blazar variability. In this presentation, the first joint study of light curves from the two instruments will be shown, correlating long-term measurements with daily sampling between air and water Cherenkov telescopes. The presented results focus on the study of the variability of the bright blazars Mrk 421 and Mrk 501 during the last two years featuring various flaring activities.Comment: 6 pages, 2 figures. Contribution to the 6th International Symposium on High Energy Gamma-Ray Astronomy (Gamma2016), Heidelberg, Germany. To be published in the AIP Conference Proceeding

Contribución al conocimiento de Porosagrotis gypaetina (Guen.) (Lep.:Noctuidae)

p.15-22Este trabajo tiene por finalidad brindar una descripcion detallada de los diferentes estados de desarrollo, asi como de los estadios larvales, de Porosagrotis gypaetina (Guen.) y estimar sus principales parametros biologicos. Se trata de una oruga conocida vulgarmente como gusano pardo que frecuenta cultivos de alfalfa, trebol bianco, maiz y girasol y determinadas malezas. Los caracteres considerados para su identificacion fueron, en el huevo: numero y distribucion de costas; en la larva: pigmentacion, distribucion de manchas y cerdas corporales; en la pupa: tamaño, forma y color y caracteristicas del cremaster; y en el adulto: ubicacion y coloracion de maculas y nervaduras alares. La emergencia de imagos alcanzo su maximo en abril y mayo. El periodo embrionario se completo en 22 a 26 dias. Aproximadamente la mitad de las larvas cumplieron su ciclo en 6 estadios y las restantes en 7; la duracion total del periodo larval fue de 134 a 141 dias, sin considerar la forma prepupal e independientemente del numero de estadios. Las orugas permanecieron como prepupas durante la temporada estival (aproximadamente 161 dias). El estado pupal duro 40 a 57 dias. Las observaciones realizadas permiten expresar que, inediante los caracteres descriptos, es factible reconocer la especie a traves no solo de los adultos, sino de sus estados inmaduros. Posee una sola generacion anual; transcurre el inviemo como larva; el daño tipico de corte lo produce a partir del cuarto estadio larval

Long-term monitoring of bright blazars in the multi-GeV to TeV range with FACT

Blazars like Markarian 421 or Markarian 501 are active galactic nuclei (AGN), with their jets orientated towards the observer. They are among the brightest objects in the very high energy (VHE) gamma ray regime (>100 GeV). Their emitted gamma-ray fluxes are extremely variable, with changing activity levels on timescales between minutes, months, and even years. Several questions are part of the current research, such as the question of the emission regions or the engine of the AGN and the particle acceleration. A dedicated longterm monitoring program is necessary to investigate the properties of blazars in detail. A densely sampled and unbiased light curve allows for observation of both high and low states of the sources, and the combination with multi-wavelength observation could contribute to the answer of several questions mentioned above. FACT (First G-APD Cherenkov Telescope) is the first operational telescope using silicon photomultiplier (SiPM, also known as Geigermode—Avalanche Photo Diode, G-APD) as photon detectors. SiPM have a very homogenous and stable longterm performance, and allow operation even during full moon without any filter, leading to a maximal duty cycle for an Imaging Air Cherenkov Telescope (IACT). Hence, FACT is an ideal device for such a longterm monitoring of bright blazars. A small set of sources (e.g., Markarian 421, Markarian 501, 1ES 1959+650, and 1ES 2344+51.4) is currently being monitored. In this contribution, the FACT telescope and the concept of longterm monitoring of bright blazars will be introduced. The results of the monitoring program will be shown, and the advantages of densely sampled and unbiased light curves will be discussed

New findings at the morphological characterisation of enteric coinnervation on esophageal striated muscle fibres.

Hintergrund und Ziele Ein Schwerpunkt der Arbeit stellt die weitere Untersuchung der enterischer Coinnervation an quergestreifter Muskulatur der Speiseröhre dar. Dieses spezielle Innervationsschema wurde an verschiedenen Tierarten sowie beim Menschen nachgewiesen. So zeigen phylogenetisch alte Säuger wie die Fledermaus, aber auch Wiederkäuer wie Antilope und Schraubenziege, enterische Coinnervation. In dieser Arbeit vervollständigten wir die Tiergruppen, indem zusätzlich Speiseröhren von Spitzmaus und Schaf untersucht und quantifiziert wurden. Da es im Gegensatz zu pharmakologischen und funktionellen Untersuchungen kaum morphologische Daten zu Serotonin (5-HT) in der Speiseröhre gibt, fokussiert diese Arbeit vor allem auch auf dem Nachweis von Serotonin in der Speiseröhre der Maus. Antikörper gegen verschiedene neuronale Marker (vasoaktives intestinales Peptid (VIP), Cholinazetyltransferase (ChAT), Tyrosinhydroxylase (TH)) wurden benutzt, um festzustellen, mit welchen Markern Serotonin möglicherweise colokalisiert ist. Weiterhin galt es zu klären, ob Serotonin tatsächlich in enterischen Neuronen produziert oder aus externen Quellen aufgenommen wird. Methoden Die Untersuchungen wurden an 76 adulten und drei juvenilen C57/Bl6 Mäusen, einem Schaf und drei Spitzmäusen durchgeführt. Außerdem wurde Gewebe von 12 Tryptophanhydroxylase (TPH)-Knockout-Mäusen untersucht. Bei einem Teil der Mäuse wurde das Material unfixiert entnommen, in begaster Krebs-Ringer-Lösung inkubiert und anschließend mit 4 % Paraformaldehyd (PFA) immersionsfixiert. Ein Teil der Mäuse wurde mit 4 % PFA perfusionsfixiert und anschließend das Gewebe entnommen. An Gefrierschnitten von Speiseröhre, Dünndarm und Gehirn sowie an Häutchenpräparaten von Speiseröhre und Dünndarm wurden sowohl histologische als auch enzym- bzw. immunhistochemische Färbungen durchgeführt, ausgewertet und dokumentiert. Ergebnisse Enterische Coinnervation der quergestreiften Muskulatur der Speiseröhre konnte bei Spitzmaus und Schaf nachgewiesen werden. 5-HT-positive Nervenfasern konnten in der Lamina muscularis mucosae, der Tunica muscularis, im myenterischen Plexus, sowie um Blutgefäße nachgewiesen werden. Des Weiteren konnten wir stark 5-HT-immunoreaktive Mastzellen in der Tela submucosa und Tunica adventitia darstellen. Etwa 10 % der enterischen Neurone waren 5-HT-positiv. Etwa 13 % aller motorischen Endplatten wurden von 5-HT-positiven Nervenendigungen kontaktiert. Die Anzahl der 5-HT-positiven Neurone, die mit VIP colokalisiert waren lag bei ca. 30 %, mit ChAT zeigten lediglich 3 % und mit TH 6 % der 5-HT-positiven Neurone Colokalisation. An motorischen Endplatten waren ca. 5 % aller 5-HT-positiven Nervenfasern auch VIP-immunreaktiv sowie 64 % TH-positiv. Im Nucleus ambiguus waren keine Nervenzellkörper positiv für 5-HT, jedoch zahlreiche Nervenfasern, welche vermutlich aus den 5-HT-positiven Neuronen der Raphekerne stammen. Ob Serotonin in enterischen Neuronen der Speiseröhre produziert wird, konnte nicht eindeutig nachgewiesen werden. Antikörper gegen TPH zeigten keine zufriedenstellende Färbung bzw. reagierten kreuz mit TH. Selbst Untersuchungen an TPH-Knockout-Mäuse lieferten kein eindeutiges Ergebnis. Schlussfolgerung Kenntnisse über das Auftreten enterischer Coinnervation bei unterschiedlichen Spezies und Serotonin als neuen und weiteren Transmitter dieses Innervationsschemas liefern uns weitere wertvolle Informationen über das funktionelle Ausmaß enterischer Modulation der quergestreiften Ösophagusmuskulatur. Serotonin dürfte die quergestreifte Speiseröhrenmuskulatur nicht nur auf zentraler Ebene via Projektionen der Raphekerne zum Nucleus ambiguus, sondern auch peripher via enterischer Coinnervation beeinflussen. Weiterhin liefern wir mit der Charakterisierung von Serotonin in der Speiseröhre der Maus neue Aspekte, welche bisherige pharmakologische und funktionelle Daten besser interpretieren helfen. Ob Serotonin in der Speiseröhre tatsächlich intrinsischen Ursprungs ist, konnte nicht abschließend geklärt werden. Zumindest ein Teil der 5-HT-positiven Nervenfasern in der Speiseröhre dürfte jedoch sympathischer Herkunft sein.Background and Aim In the present work, we focused on enteric coinnervation of striated muscle fibers in the esophagus. Enteric coinnervation has been demonstrated in several species including human. For example, the esophagus of phylogenetically old mammals like the bat and also ruminants like the antelope and he-goat show this innervation scheme. This work was undertaken to investigate esophagi of sheep and shrew to complete the category of ruminants and old mammals. As little is known about the morphological organization of serotoninergic neurons and other serotonin-containing elements in the esophagus, the second aim of this study was to characterize the serotonin (5-HT) system in the mouse esophagus. Several neuronal markers, e.g. vasoactive intestinal peptide (VIP), choline acetyltransferase (ChAT) and tyrosine hydroxylase (TH) were tested to clarify colocalization with serotonin. In addition, the issue of intrinsic versus extrinsic origin of 5-HT was investigated. Methods Experiments were performed on 76 adult and three juvenile mice (C57/Bl6), one sheep and three shrews. Furthermore, 12 tryptophan hydroxylase (TPH)-knockout-mice were included. The tissues of some mice was excised, incubated in oxygenated Krebs-Ringer solution, and subsequently fixed in 4 % phosphate-buffered formaldehyde. Other mice were perfused with 4 % phosphate-buffered formaldehyde before organ excision without preincubation in Krebs-Ringer solution. Cryosections were performed of esophagi, small intestine and brain. Whole-mounts were prepared of esophagi and small intestine. All samples were investigated histologically and by enzyme- and immunohistochemistry. Results We could demonstrate enteric coinnervation of striated muscle fibers in the esophagus of sheep and shrew. 5-HT-positive nerve fibers were present in the lamina muscularis mucosae, tunica muscularis, in the myenteric plexus and around blood vessels. Mast-cells in the tela submucosa und tunica adventitia were also positive for 5-HT. On average, about 10 % of all myenteric neurons were stained for 5-HT and about 13 % of all motor endplates were co-innervated by enteric nerve terminals positive for 5-HT. We found about 30 % 5-HT/VIP cell bodies, 3 % 5-HT/ChAT and 6 % 5-HT/TH neurons in the myenteric plexus. On motor endplates, we detected about 5 % 5-HT/VIP- and 64 % 5-HT/TH-positive nerve fibers. Nerve cell bodies of the nucleus ambiguus were negative for 5-HT, but were endowed with 5-HT-positive nerve fibers. These fibers probably derive from 5-HT-positive neurons of the raphe nuclei. The question, whether serotonin is also produced in enteric neurons could not definitely be resolved. Antibodies against TPH did not show adequate staining and even cross reactivity with TH. Investigation of TPH-knockout-mice provided ambiguous results. Conclusion This study provides further information about the occurrence and potential function of enteric coinnervation of esophageal striated muscle fibers. We identified enteric coinnervation in the esophagus of additional species and established serotonin as a new component in this innervation pattern. Serotonin might modulate vagal innervation of esophageal striated muscles both at the central level via projections of raphe nuclei to the compact formation of the nucleus ambiguus and at the peripheral level via enteric coinnervation of motor endplates. The characterization of serotonin in the esophagus helps to interpret previous pharmacological and functional data. However, we were not able to definitely clarify the question, whether serotonin is a genuine product of myenteric neurons. At least some 5-HT-positive nerve fibers in the esophagus may be in fact of sympathetic origin

Premature ventricular complexes and development of heart failure in a community-based population

ObjectiveA higher premature ventricular complex (PVC) frequency is associated with incident congestive heart failure (CHF) and death. While certain PVC characteristics may contribute to that risk, the current literature stems from patients in medical settings and is therefore prone to referral bias. This study aims to identify PVC characteristics associated with incident CHF in a community-based setting.MethodsThe Cardiovascular Health Study is a cohort of community-dwelling individuals who underwent prospective evaluation and follow-up. We analysed 24-hour Holter data to assess PVC characteristics and used multivariable logistic and Cox proportional hazards models to identify predictors of a left ventricular ejection fraction (LVEF) decline and incident CHF, respectively.ResultsOf 871 analysed participants, 316 participants exhibited at least 10 PVCs during the 24-hour recording. For participants with PVCs, the average age was 72±5 years, 41% were women and 93% were white. Over a median follow-up of 11 years, 34% developed CHF. After adjusting for demographics, cardiovascular comorbidities, antiarrhythmic drug use and PVC frequency, a greater heterogeneity of the PVC coupling interval was associated with an increased risk of LVEF decline and incident CHF. Of note, neither PVC duration nor coupling interval duration exhibited a statistically significant relationship with either outcome.ConclusionsIn this first community-based study to identify Holter-based features of PVCs that are associated with LVEF reduction and incident CHF, the fact that coupling interval heterogeneity was an independent risk factor suggests that the mechanism of PVC generation may influence the risk of heart failure

FACT - Performance of the first SiPM camera

The First G-APD Cherenkov Telescope (FACT) is the first operational test of the performance of silicon photomultipliers (SiPM) in Cherenkov Astronomy. These novel photo detectors promised to be an inexpensive and robust alternative for vacuum photomultiplier tubes, but had, up to now, never been applied in an Imaging Air shower Cherenkov Telescope (IACT). For more than three years FACT has operated on La Palma, Canary Islands (Spain), with the primary aim of long-term monitoring of astrophysical sources. Stable performance of the photo detectors is crucial and therefore has been studied in great detail. Special care has been taken with regards to their temperature and over voltage dependence through implementation of a feedback method for keeping their properties stable. Several independent long term measurements were conducted to analyze and verify SiPM gain stability. Dark count spectra, which also make for an excellent self calibration mechanism, were used to study and correct for temperature dependencies. Rate scans make it possible to derive a method, for quickly finding appropriate trigger thresholds by measuring pixel currents, and thus allow for a consistent data acquisition rate. Dedicated measurements using an LED flasher are used to study the correct application of SiPM bias voltages. In this paper, the results of the long term studies will be presented and the applicability of SiPMs in IACTs for long term monitoring will be shown.SCOPUS: cp.pinfo:eu-repo/semantics/publishe

FACT - Status and experience from three years operation of the first SiPM camera

The First G-APD Cherenkov Telescope (FACT) is pioneering the usage of solid state photosensors (G-APD, also known as SiPM). The 1440 pixel camera is installed in a 9.5 m2 refurbished HEGRA telescope on the Canary Island La Palma. Physics data-taking with FACT started in October 2011, a few hours after installation of the camera. Since Summer 2012, FACT is operated remotely without the need of a data-taking crew on site. During more than three years of operation of FACT, G-APDs have proven to be very reliable. Despite operating them regularly also under very strong moonlight conditions, the GAPDs show no change in their properties or any indication for aging. This allows FACT to have a successful monitoring program of the brightest TeV blazars in the Northern hemisphere and several flare-alerts have been sent to the community. This proceeding summarizes the history and status of FACT as well as reporting the lessons learned about the usage of SiPM in a Cherenkov telescope from the construction and operation of FACT.SCOPUS: cp.pinfo:eu-repo/semantics/publishe

FACT - Novel mirror alignment using bokeh and enhancement of the VERITAS SCCAN alignment method

Imaging Air Cherenkov Telescopes, including the First G-APD Cherenkov Telescope (FACT), use segmented reflectors. These offer large and fast apertures for little resources. However, one challenge is the alignment of the mirrors to gain a sharp image. For Cherenkov telescopes, high spatial and temporal resolution is crucial to reconstruct air shower events. Therefore one has to align the individual mirror positions and orientations precisely. Alignment is difficult due to the large number of degrees of freedom and, because most techniques involve a star, has to be done during good weather nights which overlaps with observation time. We present the mirror alignment of FACT, done using two methods. Firstly, we show a new method which we call Bokeh alignment. This method is simple, cheap and can even be done during daytime. Secondly, we demonstrate an enhancement of the SCCAN method by F. Arqueros et al. and first implemented by the McGill VERITAS group. Using a second camera, our enhanced SCCAN is optimized for changing weather, changing zenith distance, and changing reference stars. Developed off site in the lab on a 1/10th scale model of FACT, both our methods resulted in a highly telescope independent procedure, e.g. both our methods run without communication to the telescope's drive. We compare alignment results by using the point spread function of star images, ray tracing simulations, and overall muon rates before and after the alignment.SCOPUS: cp.pinfo:eu-repo/semantics/publishe