Loss of Neuron Navigator 2 Impairs Brain and Cerebellar Development

Acknowledgements We thank the study family for their enthusiastic participationPeer reviewedPublisher PD

Different Host Exploitation Strategies in Two Zebra Mussel-Trematode Systems: Adjustments of Host Life History Traits

The zebra mussel is the intermediate host for two digenean trematodes, Phyllodistomum folium and Bucephalus polymorphus, infecting gills and the gonad respectively. Many gray areas exist relating to the host physiological disturbances associated with these infections, and the strategies used by these parasites to exploit their host without killing it. The aim of this study was to examine the host exploitation strategies of these trematodes and the associated host physiological disturbances. We hypothesized that these two parasite species, by infecting two different organs (gills or gonads), do not induce the same physiological changes. Four cellular responses (lysosomal and peroxisomal defence systems, lipidic peroxidation and lipidic reserves) in the host digestive gland were studied by histochemistry and stereology, as well as the energetic reserves available in gonads. Moreover, two indices were calculated related to the reproductive status and the physiological condition of the organisms. Both parasites induced adjustments of zebra mussel life history traits. The host-exploitation strategy adopted by P. folium would occur during a short-term period due to gill deformation, and could be defined as “virulent.” Moreover, this parasite had significant host gender-dependent effects: infected males displayed a slowed-down metabolism and energetic reserves more allocated to growth, whereas females displayed better defences and would allocate more energy to reproduction and maintenance. In contrast, B. polymorphus would be a more “prudent” parasite, exploiting its host during a long-term period through the consumption of reserves allocated to reproduction

Drosophila Voltage-Gated Sodium Channels Are Only Expressed in Active Neurons and Are Localized to Distal Axonal Initial Segment-like Domains

In multipolar vertebrate neurons, action potentials (APs) initiate close to the soma, at the axonal initial segment. Invertebrate neurons are typically unipolar with dendrites integrating directly into the axon. Where APs are initiated in the axons of invertebrate neurons is unclear. Voltage-gated sodium (NaV) channels are a functional hallmark of the axonal initial segment in vertebrates. We used an intronic Minos-Mediated Integration Cassette to determine the endogenous gene expression and subcellular localization of the sole NaV channel in both male and female Drosophila, para Despite being the only NaV channel in the fly, we show that only 23 ± 1% of neurons in the embryonic and larval CNS express para, while in the adult CNS para is broadly expressed. We generated a single-cell transcriptomic atlas of the whole third instar larval brain to identify para expressing neurons and show that it positively correlates with markers of differentiated, actively firing neurons. Therefore, only 23 ± 1% of larval neurons may be capable of firing NaV-dependent APs. We then show that Para is enriched in an axonal segment, distal to the site of dendritic integration into the axon, which we named the distal axonal segment (DAS). The DAS is present in multiple neuron classes in both the third instar larval and adult CNS. Whole cell patch clamp electrophysiological recordings of adult CNS fly neurons are consistent with the interpretation that Nav-dependent APs originate in the DAS. Identification of the distal NaV localization in fly neurons will enable more accurate interpretation of electrophysiological recordings in invertebrates.SIGNIFICANCE STATEMENT The site of action potential (AP) initiation in invertebrates is unknown. We tagged the sole voltage-gated sodium (NaV) channel in the fly, para, and identified that Para is enriched at a distal axonal segment. The distal axonal segment is located distal to where dendrites impinge on axons and is the likely site of AP initiation. Understanding where APs are initiated improves our ability to model neuronal activity and our interpretation of electrophysiological data. Additionally, para is only expressed in 23 ± 1% of third instar larval neurons but is broadly expressed in adults. Single-cell RNA sequencing of the third instar larval brain shows that para expression correlates with the expression of active, differentiated neuronal markers. Therefore, only 23 ± 1% of third instar larval neurons may be able to actively fire NaV-dependent APs

Drosophila Voltage-Gated Sodium Channels Are Only Expressed in Active Neurons and Are Localized to Distal Axonal Initial Segment-like Domains

In multipolar vertebrate neurons, action potentials (APs) initiate close to the soma, at the axonal initial segment. Invertebrate neurons are typically unipolar with dendrites integrating directly into the axon. Where APs are initiated in the axons of invertebrate neurons is unclear. Voltage-gated sodium (NaV) channels are a functional hallmark of the axonal initial segment in vertebrates. We used an intronic Minos-Mediated Integration Cassette to determine the endogenous gene expression and subcellular localization of the sole NaV channel in both male and female Drosophila, para Despite being the only NaV channel in the fly, we show that only 23 ± 1% of neurons in the embryonic and larval CNS express para, while in the adult CNS para is broadly expressed. We generated a single-cell transcriptomic atlas of the whole third instar larval brain to identify para expressing neurons and show that it positively correlates with markers of differentiated, actively firing neurons. Therefore, only 23 ± 1% of larval neurons may be capable of firing NaV-dependent APs. We then show that Para is enriched in an axonal segment, distal to the site of dendritic integration into the axon, which we named the distal axonal segment (DAS). The DAS is present in multiple neuron classes in both the third instar larval and adult CNS. Whole cell patch clamp electrophysiological recordings of adult CNS fly neurons are consistent with the interpretation that Nav-dependent APs originate in the DAS. Identification of the distal NaV localization in fly neurons will enable more accurate interpretation of electrophysiological recordings in invertebrates.SIGNIFICANCE STATEMENT The site of action potential (AP) initiation in invertebrates is unknown. We tagged the sole voltage-gated sodium (NaV) channel in the fly, para, and identified that Para is enriched at a distal axonal segment. The distal axonal segment is located distal to where dendrites impinge on axons and is the likely site of AP initiation. Understanding where APs are initiated improves our ability to model neuronal activity and our interpretation of electrophysiological data. Additionally, para is only expressed in 23 ± 1% of third instar larval neurons but is broadly expressed in adults. Single-cell RNA sequencing of the third instar larval brain shows that para expression correlates with the expression of active, differentiated neuronal markers. Therefore, only 23 ± 1% of third instar larval neurons may be able to actively fire NaV-dependent APs.status: publishe

Dopaminergic transmission in striatal slices of Pink1 WT and KO mice.

<p><b>A</b> Scatter plots illustrating the values of the peak [DA] (left panel) and the half-life (right panel) measured from averaged single-pulse transients. Bars represent mean and SEM, no statistical differences were detected when comparing the WT and KO genotypes (two-tailed unpaired t test p = 0.995 and p = 0.556 DA peak and half-life, respectively). <b>B</b> Time course of the normalized peak of the evoked responses to single pulses (black arrows) double pulses (gray arrows) and trains (red arrows). Genotypes were compared by means of a 2 Way Repeated Measures ANOVA yielding no significant difference (p = 0.2100; n = 12 WT and n = 12 KO). The green box indicates the presence of 5 µM nomifensine into circulating ACSF.</p

Dopaminergic transmission evaluated using FSCV.

<p>Electrically evoked DA transients were detected by FSCV in the dorso-lateral region of slices containing the most rostral part of the striatum. <b>A</b> Heat-map plot showing the measured current as a function of time and applied potential. <b>B</b> Extracellular DA overflow measured by the carbon fiber electrode at a sampling rate of 10 Hz (black trace) and the exponential fit (red trace) applied from the peak to the end of the signal (half-life: 0.2723 s, 95% confidence intervals: 0.2633 to 0.2821 s; R²: 0.9646); arrow-head represents the time of stimulation. Inset: voltammogram corresponding to the peak of DA concentration, depicting maximal oxidation current at a potential of 306.12 mV (vertical dotted line). <b>C</b> Averaged 5 consecutive responses to either single-pulse, double pulse or train stimulation in normal ACSF (left) or in the presence of 5 µM nomifensine. <b>D</b> Scatter plots representing the peak DA concentration elicited by the different stimuli. Significant differences were found between the first single and double pulse stimulation in normal ACSF and between the fifth single pulse and the first train (p = 0.0010; p = 0.0020, respectively. Two-tailed Wilcoxon matched pairs test). In the presence of 5 µM nomifensine both comparisons yielded significant differences (both p = 0.0010. Two-tailed Wilcoxon matched pairs-test). All data presented in this figure were generated from experiments with WT animals belonging to the Parkin colony (slices n = 11; animals n = 11).</p

Dopaminergic transmission in striatal slices of WT and LRRK2(R1441G) transgenic mice.

<p><b>A</b> Scatter plots illustrating the values of the peak [DA] (left panel) and the half-life (right panel) measured from averaged single-pulse transients. Bars represent mean and SEM, no statistical differences were detected when comparing the WT and transgenic genotypes (two-tailed unpaired t test p  = 0.5421 and p = 0.7671 DA peak and half-life, respectively). <b>B</b> Time course of the normalized peak of the evoked responses to single pulses (black arrows) double pulses (gray arrows) and trains (red arrows). Genotypes were compared by means of a 2 Way Repeated Measures ANOVA yielding no significant difference (p = 0.4337; n = 11 WT and n = 13 Tg, animals n = 10 each). The green box indicates the presence of 5 µM nomifensine into circulating ACSF.</p