Gcamp

GCaMP is a genetically encoded calcium indicator (GECI) widely used in neuroscience research. It measures intracellular Ca2+ level by fluorescence changes as it directly binds to Ca2+ . In this process, the effect of this calcium buffer on the intracellular calcium signaling and cell physiology is often not taken into consideration. However, growing evidence from calcium imaging studies shows GCaMP expression under certain conditions can generate aberrant activity, such as seizures. In this study, we examined the effect of GCaMP6 expression in the dentate gyrus (DG) on epileptogenesis...

SIGNIFICANCE: Genetically encoded calcium ion (<mml:math xmlns:mml="https://www.w3.org/1998/Math/MathML"><mml:mrow><mml:msup><mml:mi>Ca</mml:mi><mml:mrow><mml:mn>2</mml:mn><mml:mo>+</mml:mo></mml:mrow></mml:msup></mml:mrow></mml:math>) indicators (GECIs) are powerful tools for monitoring intracellular <mml:math xmlns:mml="https://www.w3.org/1998/Math/MathML"><mml:mrow><mml:msup><mml:mi>Ca</mml:mi><mml:mrow><mml:mn>2</mml:mn><mml:mo>+</mml:mo></mml:mrow></mml:msup></mml:mrow></mml:math> concentration changes in living cells and model organisms...

The default mode network (DMN) is a large-scale brain network known to be suppressed during a wide range of cognitive tasks. However, our comprehension of its role in naturalistic and unconstrained behaviors has remained elusive because most research on the DMN has been conducted within the restrictive confines of MRI scanners. Here, we use multisite GCaMP (a genetically encoded calcium indicator) fiber photometry with simultaneous videography to probe DMN function in awake, freely exploring rats. We examined neural dynamics in three core DMN nodes-the retrosplenial cortex, cingulate cortex, and prelimbic cortex-as well as the anterior insula node of the salience network, and their association with the rats' spatial exploration behaviors...

Here we used machine learning to engineer genetically encoded fluorescent indicators, protein-based sensors critical for real-time monitoring of biological activity. We used machine learning to predict the outcomes of sensor mutagenesis by analyzing established libraries that link sensor sequences to functions. Using the GCaMP calcium indicator as a scaffold, we developed an ensemble of three regression models trained on experimentally derived GCaMP mutation libraries. The trained ensemble performed an in silico functional screen on 1,423 novel, uncharacterized GCaMP variants...

SIGNIFICANCE: The development of imaging systems that are cost-efficient and modular is essential for modern neuroscience research. AIM: In the current study, we designed, developed, and characterized a low-cost reversible tandem lens mesoscope for brain imaging in rodents. APPROACH: Using readily available components, we assembled a robust imaging system that is highly efficient and cost-effective. We developed a mesoscope that offers high-resolution structural and functional imaging with cost-effective lenses and CMOS camera...

Loss of dopamine neurons causes motor deterioration in Parkinson's disease patients. We have previously reported that in addition to acute motor impairment, the impaired motor behavior is encoded into long-term memory in an experience-dependent and task-specific manner, a phenomenon we refer to as aberrant inhibitory motor learning. Although normal motor learning and aberrant inhibitory learning oppose each other and this is manifested in apparent motor performance, in the present study, we found that normal motor memory acquired prior to aberrant inhibitory learning remains preserved in the brain, suggesting the existence of independent storage...

The ability to record every spike from every neuron in a behaving animal is one of the holy grails of neuroscience. Here, we report coming one step closer towards this goal with the development of an end-to-end pipeline that automatically tracks and extracts calcium signals from individual neurons in the cnidarian Hydra vulgaris. We imaged dually labeled (nuclear tdTomato and cytoplasmic GCaMP7s) transgenic Hydra and developed an open-source Python platform (TraSE-IN) for the Tracking and Spike Estimation of Individual Neurons in the animal during behavior...

The endoplasmic reticulum (ER) is an important regulator of Ca 2+ in cells and dysregulation of ER calcium homeostasis can lead to numerous pathologies. Understanding how various pharmacological and genetic perturbations of ER Ca 2+ homeostasis impacts cellular physiology would likely be facilitated by more quantitative measurements of ER Ca 2+ levels that allow easier comparisons across conditions. Here, we developed a ratiometric version of our original ER-GCaMP probe that allows for more quantitative comparisons of the concentration of Ca 2+ in the ER across cell types and sub-cellular compartments...

Obesity results from excessive caloric input associated with overeating and presents a major public health challenge. The hypothalamus has received significant attention for its role in governing feeding behavior and body weight homeostasis. However, extra-hypothalamic brain circuits also regulate appetite and consumption by altering sensory perception, motivation, and reward. We recently discovered a population of basal forebrain cholinergic (BFc) neurons that regulate appetite suppression. Through viral tracing methods in the mouse model, we found that BFc neurons densely innervate the basolateral amygdala (BLA), a limbic structure involved in motivated behaviors...

Lanthanides, a series of 15 f-block elements, are crucial in modern technology, and their purification by conventional chemical means comes at a significant environmental cost. Synthetic biology offers promising solutions. However, progress in developing synthetic biology approaches is bottlenecked because it is challenging to measure lanthanide binding with current biochemical tools. Here we introduce LanTERN, a lanthanide-responsive fluorescent protein. LanTERN was designed based on GCaMP, a genetically encoded calcium indicator that couples the ion binding of four EF hand motifs to increased GFP fluorescence...

Even under spontaneous conditions and in the absence of changing environmental demands, awake animals alternate between increased or decreased periods of alertness. These changes in brain state can occur rapidly, on a timescale of seconds, and neuromodulators such as acetylcholine (ACh) are thought to play an important role in driving these spontaneous state transitions. Here, we perform the first simultaneous imaging of ACh sensors and GCaMP-expressing axons in vivo , to examine the spatiotemporal properties of cortical ACh activity and release during spontaneous changes in behavioral state...

One goal of neurobiology is to explain how decision-making in neuromuscular circuits produces behaviors. However, two obstacles complicate such efforts: individual behavioral variability and the challenge of simultaneously assessing multiple neuronal activities during behavior. Here, we circumvent these obstacles by analyzing whole animal behavior from a library of Caenorhabditis elegans male mating recordings. The copulating males express the GCaMP calcium sensor in the muscles, allowing simultaneous recording of posture and muscle activities...

Temporomandibular disorder (TMD) is the most prevalent painful condition in the craniofacial area. The pathophysiology of TMD is not fully understood, and it is necessary to understand pathophysiology underlying painful TMD conditions to develop more effective treatment methods. Recent studies suggested that external or intrinsic trauma to TMJ is associated with chronic TMD in patients. Here, we investigated the effects of the TMJ trauma through forced-mouth opening (FMO) in mice to determine pain behaviors and peripheral sensitization of trigeminal nociceptors...

Patients with temporomandibular disorders (TMD) typically experience facial pain and discomfort or tenderness in the temporomandibular joint (TMJ), causing disability in daily life. Unfortunately, existing treatments for TMD are not always effective, creating a need for more advanced, mechanism-based therapies. In this study, we used in vivo GCaMP3 Ca 2+ imaging of intact trigeminal ganglia (TG) to characterize functional activity of the TG neurons in vivo , specifically in TMJ animal models. This system allows us to observe neuronal activity in intact anatomical, physiological, and clinical conditions and to assess neuronal function and response to various stimuli...

The arcuate nucleus kisspeptin (ARNKISS) neurons represent the gonadotropin-releasing hormone (GnRH) pulse generator that likely drives pulsatile gonadotropin secretion in all mammals. Using an improved GCaMP fiber photometry system enabling long-term continuous recordings, we aimed here to establish a definitive profile of ARNKISS neuronal activity across the murine estrous cycle. As noted previously, a very substantial reduction in the frequency of ARNKISS neuron synchronization events (SEs) occurs on late proestrus and extends into estrus...

Head-mounted miniscopes have allowed for functional fluorescence imaging in freely moving animals. However, current capabilities of state-of-the-art technology can record only up to two, spectrally distinct fluorophores. This severely limits the number of cell types identifiable in a functional imaging experiment. Here we present a pipeline that enables the distinction of nine neuronal subtypes from regions defined by behaviorally relevant cells during in vivo GCaMP imaging. These subtypes are identified utilizing unique fluorophores that are co-expressed with GCaMP, unmixed by spectral imaging on a confocal microscope and co-registering these spectral fingerprints with functional data obtained on miniaturized microscopes...

Viral expression of the calcium indicator GCaMP in primate RGCs has enabled optical readout of retinal function at a cellular scale in vivo. To date, functional recording has been limited to transduced RGCs close to the foveal pit. In this study we evaluate ILM peel as a strategy to expand the area of transduced RGCs and allow functional recording beyond the fovea in the living eye. 4 eyes of 3 immunosuppressed macaca fascicularis received a 9-12° ILM peel centered on the fovea, followed by intravitreal injection of GCaMP8s 4-8 weeks post-peel...

UNLABELLED: To survive daily damage, the formation of actomyosin ring at the wound periphery is required to rapidly close cell wounds. Calcium influx is one of the start signals for these cell wound repair events. Here, we find that rapid recruitment of all three Drosophila calcium responding and phospholipid binding Annexin proteins (AnxB9, AnxB10, AnxB11) to distinct regions around the wound are regulated by the quantity of calcium influx rather than their binding to specific phospholipids...

We present a cost-effective, compact foot-print, and open source Raspberry Pi-based wide field imaging system. The compact nature allows the system to be used for close-proximity dual-brain cortical mesoscale functional-imaging to simultaneously observe activity in two head-fixed animals in a staged social touch-like interaction. We provide all schematics, code, and protocols for a rail system where head-fixed mice are brought together to a distance where the macrovibrissae of each mouse make contact. Cortical neuronal functional signals (GCaMP6s; genetically encoded Ca2+ sensor) were recorded from both mice simultaneously before, during, and after the social contact period...

Optical methods for studying the brain are powerful tools for understanding how neural activity underlies complex behavior. These methods typically rely on genetically encoded sensors and actuators to monitor and control neural activity. For microendoscopic calcium imaging, injection of a virus followed by implantation of a lens probe is required to express a calcium sensor and enable optical access to the target brain region. This two-step process poses several challenges, chief among them being the risks associated with mistargeting and/or misalignment between virus expression zone, lens probe and target brain region...