Neurites

Patients with densely innervated tumors suffer with increased metastasis and decreased survival as compared to those with less innervated tumors. We hypothesize that in some tumors, nerves are acquired by a tumor-induced process, called axonogenesis. Here, we use PC12 cells as an in vitro neuronal model, human tumor samples and murine in vivo models to test this hypothesis. When appropriately stimulated, PC12 cells extend processes, called neurites. We show that patient tumors release vesicles, called exosomes, which induce PC12 neurite outgrowth. Using a cancer mouse model, we show that tumors compromised in exosome release are less innervated than controls. Moreover, in vivo pharmacological blockade of exosome release similarly attenuates tumor innervation. We characterize these nerves as sensory in nature and demonstrate that axonogenesis is potentiated by the exosome-packaged axonal guidance molecule, EphrinB1. These findings indicate that tumor released exosomes induce tumor innervation and exosomes containing EphrinB1 potentiate this activity.

The PC12 cell line is one of the most commonly used in neuroscience research, including studies on neurotoxicity, neuroprotection, neurosecretion, neuroinflammation, and synaptogenesis. Two types of this line are available in the ATCC collection: traditional PC12 cells grown in suspension and well-attached adherent phenotype. PC12 cells grown in suspension tend to aggregate and adhere poorly to non-coated surfaces. Therefore, it is necessary to modify the surface of culture vessels. This paper aims to characterise the use of two distinct variants of PC12 cells as well as describe their differentiation and neuronal outgrowth with diverse NGF concentrations (rat or human origin) on various surfaces. In our study, we evaluated cell morphology, neurite length, density and outgrowth (measured spectrofluorimetrically), and expression of neuronal biomarkers (doublecortin and NeuN). We found that the collagen coating was the most versatile method of surface modification for both cell lines. For adherent cells, the coating was definitely less important, and the poly-d-lysine surface was as good as collagen. We also demonstrated that the concentration of NGF is of great importance for the degree of differentiation of cells. For suspension cells, we achieved the best neuronal characteristics (length and density of neurites) after 14 days of incubation with 100 ng/mL NGF (change every 48 h), while for adherent cells after 3-5 days, after which they began to proliferate. In the PC12 cell line, doublecortin (DCX) expression in the cytoplasm and NeuN in the cell nucleus were found. In turn, in the PC12 Adh line, DCX was not expressed, and NeuN expression was located in the entire cell (both in the nucleus and cytoplasm). Only the traditional PC12 line grown in suspension after differentiation with NGF should be used for neurobiological studies, especially until the role of the NeuN protein, whose expression has also been noted in the cytoplasm of adherent cells, is well understood.

Neurons undergo substantial morphological and functional changes during development to form precise synaptic connections and acquire specific physiological properties. What are the underlying transcriptomic bases? Here, we obtained the single-cell transcriptomes of Drosophila olfactory projection neurons (PNs) at four developmental stages. We decoded the identity of 21 transcriptomic clusters corresponding to 20 PN types and developed methods to match transcriptomic clusters representing the same PN type across development. We discovered that PN transcriptomes reflect unique biological processes unfolding at each stage-neurite growth and pruning during metamorphosis at an early pupal stage; peaked transcriptomic diversity during olfactory circuit assembly at mid-pupal stages; and neuronal signaling in adults. At early developmental stages, PN types with adjacent birth order share similar transcriptomes. Together, our work reveals principles of cellular diversity during brain development and provides a resource for future studies of neural development in PNs and other neuronal types.

The type C variant (TDP-C) of FTLD-TDP exhibits unique features, not shared by types A and B, namely the invariable and frequently asymmetric predilection for the anterior temporal lobes (ATL). Depending on the direction of hemispheric asymmetry, the associated clinical features include word comprehension impairment, associative agnosia, and behavioral abnormalities. Current research on TDP-C aims to explore the factors that underlie the selective targeting of the ATL and, more specifically, the cellular details that undermine the behavioral and cognitive functions of this region. Abnormal TDP-C neurites have recently been shown to represent heterodimers with annexin A11 (ANXA11). This feature, not shared by TDP-A or -B, may explain the unique predilection of TDP-C for the ATL. To further explore the subcellular distribution of the pathology, paraffin-embedded sections were stained using fluorescent antibodies for the dendritic marker MAP2 and phosphorylated TDP-43 (pTDP) or ANXA11. Results indicated that approximately half of pTDP/ANXA11 neurites co-localized with MAP2. The actual overlap during life may be much higher but decreased at autopsy through dendritic loss due to prolonged neurodegeneration. The potentially selective and progressive dendritic pathology of TDP-C, quite unique among neurodegenerative entities, may underlie the distinctive perturbation of cortical integrative computations.