Electrophoresis of polar fluorescent tracers through the nerve sheath labels neuronal populations for anatomical and functional imaging

Isaacson, M.; Hedwig, Berthold

doi:10.1038/srep40433

Cited by 6 publications

(5 citation statements)

References 17 publications

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“…removing the cuticle). Additionally, the ability to image through the cuticle allows an alternative approach to traditional electrophysiology, such as the use of voltage-sensitive dyes to follow neuron activity in real time of the mechano-sensory cells involved in the hearing process [42][43][44]. Furthermore, from the point of view of invasive experimental protocols, invertebrates, and especially insects, are ideal substitutes within the 3Rs framework [45].…”

Section: Discussionmentioning

confidence: 99%

Non-invasive biophysical measurement of travelling waves in the insect inner ear

et al. 2017

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Frequency analysis in the mammalian cochlea depends on the propagation of frequency information in the form of a travelling wave (TW) across tonotopically arranged auditory sensilla. TWs have been directly observed in the basilar papilla of birds and the ears of bush-crickets (Insecta: Orthoptera) and have also been indirectly inferred in the hearing organs of some reptiles and frogs. Existing experimental approaches to measure TW function in tetrapods and bush-crickets are inherently invasive, compromising the fine-scale mechanics of each system. Located in the forelegs, the bush-cricket ear exhibits outer, middle and inner components; the inner ear containing tonotopically arranged auditory sensilla within a fluid-filled cavity, and externally protected by the leg cuticle. Here, we report bush-crickets with transparent ear cuticles as potential model species for direct, non-invasive measuring of TWs and tonotopy. Using laser Doppler vibrometry and spectroscopy, we show that increased transmittance of light through the ear cuticle allows for effective non-invasive measurements of TWs and frequency mapping. More transparent cuticles allow several properties of TWs to be precisely recovered and measured in vivo from intact specimens. Our approach provides an innovative, non-invasive alternative to measure the natural motion of the sensilla-bearing surface embedded in the intact inner ear fluid.

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Section: Discussionmentioning

confidence: 99%

Non-invasive biophysical measurement of travelling waves in the insect inner ear

et al. 2017

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“…So far, calcium imaging of locust neurons required intracellular injection of calcium indicators 30 , 44 , 50 , making simultaneous calcium imaging of large cell populations difficult. However, Isaacson and Hedwig described an innovative technique, in which calcium indicators were introduced via iontophoresis through the nerve sheath to multiple cells 51 . This method appears technically feasible for easily accessible nerves, but may be rather difficult to apply for loading neurons of tracts running in the depth of the brain, such as the antennal lobe tract.…”

Section: Discussionmentioning

confidence: 99%

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

Bergmann¹,

Bicker²

2021

Sci Rep

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Locusts are advantageous organisms to elucidate mechanisms of olfactory coding at the systems level. Sensory input is provided by the olfactory receptor neurons of the antenna, which send their axons into the antennal lobe. So far, cellular properties of neurons isolated from the circuitry of the olfactory system, such as transmitter-induced calcium responses, have not been studied. Biochemical and immunocytochemical investigations have provided evidence for acetylcholine as classical transmitter of olfactory receptor neurons. Here, we characterize cell cultured projection and local interneurons of the antennal lobe by cytosolic calcium imaging to cholinergic stimulation. We bulk loaded the indicator dye Cal-520 AM in dissociated culture and recorded calcium transients after applying cholinergic agonists and antagonists. The majority of projection and local neurons respond with increases in calcium levels to activation of both nicotinic and muscarinic receptors. In local interneurons, we reveal interactions lasting over minutes between intracellular signaling pathways, mediated by muscarinic and nicotinic receptor stimulation. The present investigation is pioneer in showing that Cal-520 AM readily loads Locusta migratoria neurons, making it a valuable tool for future research in locust neurophysiology, neuropharmacology, and neurodevelopment.

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“…Suction electrode recordings were obtained from the auditory nerves of five adult C. gorgonensis following previously described methods ( Isaacson and Hedwig, 2017 ). Briefly, animals were restrained dorsal side up in plasticine with their acoustic spiracles and tympana exposed to the air.…”

Section: Methodsmentioning

confidence: 99%

Ear pinnae in a neotropical katydid (Orthoptera: Tettigoniidae) function as ultrasound guides for bat detection

Pulver

Celiker

Woodrow

et al. 2022

eLife

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Early predator detection is a key component of the predator-prey arms race and has driven the evolution of multiple animal hearing systems. Katydids (Insecta) have sophisticated ears, each consisting of paired tympana on each foreleg that receive sound both externally, through the air, and internally via a narrowing ear canal running through the leg from an acoustic spiracle on the thorax. These ears are pressure-time difference receivers capable of sensitive and accurate directional hearing across a wide frequency range. Many katydid species have cuticular pinnae which form cavities around the outer tympanal surfaces, but their function is unknown. We investigated pinnal function in the katydid Copiphora gorgonensis by combining experimental biophysics and numerical modelling using 3D ear geometries. We found that the pinnae in C. gorgonensis do not assist in directional hearing for conspecific call frequencies, but instead act as ultrasound detectors. Pinnae induced large sound pressure gains (20–30 dB) that enhanced sound detection at high ultrasonic frequencies (>60 kHz), matching the echolocation range of co-occurring insectivorous gleaning bats. These findings were supported by behavioural and neural audiograms and pinnal cavity resonances from live specimens, and comparisons with the pinnal mechanics of sympatric katydid species, which together suggest that katydid pinnae primarily evolved for the enhanced detection of predatory bats.

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Electrophoresis of polar fluorescent tracers through the nerve sheath labels neuronal populations for anatomical and functional imaging

Cited by 6 publications

References 17 publications

Non-invasive biophysical measurement of travelling waves in the insect inner ear

Non-invasive biophysical measurement of travelling waves in the insect inner ear

Cholinergic calcium responses in cultured antennal lobe neurons of the migratory locust

Ear pinnae in a neotropical katydid (Orthoptera: Tettigoniidae) function as ultrasound guides for bat detection

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