Forward locomotion of Drosophila melanogaster larvae is composed of rhythmic waves of contractions that are thought to be produced by segmentally organized central pattern generators. We present a systematic description of spike activity patterns during locomotive contraction waves in semi-intact wild-type and mutant larval preparations. We have shown previously that Th nM18 mutants, with altered levels of octopamine and tyramine, have a locomotion deficit. By recording en passant from the segmental nerves, we investigated the coordination of the neuronal activity driving contraction waves of the abdominal body-wall muscles. Rhythmic bursts of activity that occurred concurrently with locomotive waves were frequently observed in wild-type larvae but were rarely seen in Th nM18 mutants. These centrally generated patterned activities were eliminated in the distal stumps of both wild-type and Th nM18 larvae after severing the segmental nerve from the CNS. Patterned activities persisted in the proximal stumps deprived of sensory feedback from the periphery. Simultaneous recordings demonstrated a delay in the bursting activity between different segments, with greater delay for segments that were farther apart. In contrast, bilateral recordings within a single segment revealed a well synchronized activity pattern in nerves innervating each hemisegment in both wild-type and Th nM18 larvae. Significantly, rhythmic patterns of bursts and waves could be evoked in Th nM18 mutants by head or tail stimulation despite their highly irregular spontaneous activities. These observations suggest a role of the biogenic amines in the initiation and modulation of motor pattern generation. The technique presented here can be readily extended to examine the locomotion motor program of other mutants.
Biogenic amines are believed to play important roles in producing behaviors. Although some biogenic amines have been extensively studied in both vertebrates and invertebrates, little is known about the effects of trace amines like tyramine and octopamine. We investigated how trace amines affect behaviors using quantitative morphometric methods on Drosophila Tbetah(nM18) and iav(N) mutants that have altered levels of tyramine and octopamine. Locomotion of wild-type and mutant third instar larvae was analyzed using Dynamic Image Analysis System (DIAS) software. We found that Tbetah(nM18) mutants, with elevated tyramine levels and reduced octopamine levels, had a severe locomotion phenotype. Mutant larvae spent much more time in pausing episodes than wild-type larvae and displayed a reduction in speed and linear translocation. The locomotion phenotype was partially rescued by feeding Tbetah(nM18) larvae octopamine, an effect that could be nullified with simultaneous feeding of tyramine. Feeding Tbetah(nM18) larvae yohimbine, an agent that inhibits the activity of Drosophila tyramine receptors, also improved some locomotion parameters. Feeding both octopamine and yohimbine further improved rescue efficiency. Simultaneously reducing the octopamine and tyramine levels as in iav(N) larvae, in contrast, led to a less severe behavioral phenotype than that of Tbetah(nM18) mutants. Feeding iav(N) larvae either tyramine or octopamine exerted only a minor improvement in locomotion. These results suggest that tyramine and octopamine have opposite effects on Drosophila larval locomotion regulation and that a balance between the two is important in producing normal behavior.
Under appropriate culture conditions, bone marrow (BM)-derived mesenchymal stem cells are capable of differentiating into diverse cell types unrelated to their phenotypical embryonic origin, including neural cells. Here, we report the successful generation of neural stem cell (NSC)-like cells from BM-derived human mesenchymal stem cells (hMSCs). Initially, hMSCs were cultivated in a conditioned medium of human neural stem cells. In this culture system, hMSCs were induced to become NSC-like cells, which proliferate in neurosphere-like structures and express early NSC markers. Like central nervous system-derived NSCs, these BM-derived NSC-like cells were able to differentiate into cells expressing neural markers for neurons, astrocytes, and oligodendrocytes. Whole-cell patch clamp recording revealed that neuron-like cells, differentiated from NSC-like cells, exhibited electrophysiological properties of neurons, including action potentials. Transplantation of NSC-like cells into mouse brain confirmed that these NSC-like cells retained their capability to differentiate into neuronal and glial cells in vivo. Our data show that multipotent NSC-like cells can be efficiently produced from BM-derived hMSCs in culture and that these cells may serve as a useful alternative to human neural stem cells for potential clinical applications such as autologous neuroreplacement therapies.
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