Developing neural networks follow common trends such as expression of spontaneous, recurring activity patterns, and appearance of neuromodulation. How these processes integrate to yield mature, behaviorally relevant activity patterns is largely unknown. We examined the integration of serotonergic neuromodulation and its role in the functional organization of the accessible locomotor network in developing zebrafish at behavioral and cellular levels. Locally restricted populations of serotonergic neurons and their projections appeared in the hindbrain and spinal cord of larvae after hatching (approximately day 2). However, 5-HT affected the swimming pattern only from day 4 on, when sustained spontaneous swimming appeared. 5-HT and its agonist quipazine increased motor output by reducing intervals of inactivity, observed behaviorally (by high-speed video) and in recordings from spinal neurons during fictive swimming (by whole-cell current clamp). 5-HT and quipazine had little effect on the properties of the activity periods, such as the duration of swim episodes and swim frequency. Further, neuronal input resistance, rheobasic current, and resting potential were not affected significantly. The 5-HT antagonists methysergide and ketanserin decreased motor output by prolonging the periods of inactivity with little effect on the active swim episode or neuronal properties. Our results suggest that 5-HT neuromodulation is integrated early in development of the locomotor network to increase its output by reducing periods of inactivity with little effect on the activity periods, which in contrast are the main targets of 5-HT neuromodulation in neonatal and adult preparations.

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The neurohypophysial peptide arginine vasotocin, and its mammalian ortholog arginine vasopressin, influence a wide range of physiological and behavioral responses, including aspects of sexual and social behaviors, osmoregulation, stress response, metabolism, blood pressure, and circadian rhythms. Here, we demonstrate that, in zebrafish (Danio rerio), the vasotocin precursor gene arginine vasotocin-neurophysin (avt) is expressed in two domains in the developing embryo: the dorsal preoptic area and the ventral hypothalamus. In the dorsal preoptic area, avt-expressing cells are intermingled with isotocin-neurophysin (ist) -expressing cells, and these neurons project to the neurohypophysis (posterior pituitary). In the dorsal preoptic area, the transcriptional regulators orthopedia b (otpb) and simple-minded 1 (sim1) are required for expression of both avt and ist. In contrast, otp and sim1 are not required for avt expression in the ventral hypothalamus. Thus, the development of these two avt expression domains is influenced by separate gene regulatory networks. Developmental Dynamics 237:995-1005, 2008.

The presence of galanin-like substances and their relation to substance P-, vasotocin-, and isotocin-immunoreactive neurons and fibers in the brain of teleosts was investigated with immunohistochemical methods. Two specific antisera against synthetic porcine galanin (GAL) revealed cell bodies and fibers in the brain of four different teleost species (Salmo salar, Carassius carassius, Gasterosteus aculeatus, and Anguilla anguilla). In all four species the main location of galanin immunoreactivity was in the hypothalamo-pituitary region. A detailed study of the distribution of galanin immunoreactivity in S. salar showed that galanin immunoreactive (GALir) perikarya were present in the nucleus preopticus periventricularis, an area that may be compared to the supraoptic nucleus in mammals, and in the nucleus lateralis tuberis, a nucleus involved in pituitary control in fishes that may be compared with the arcuate nucleus in mammals. GALir perikarya were found also in the nucleus recessus lateralis and in the nucleus recessus posterior. Numerous GALir fibers were present in the telencephalon and diencephalon, whereas only small numbers of fibers were found in the brainstem. In contrast to the situation in mammals, no GALir perikarya were observed in the brainstem areas corresponding to the noradrenergic locus coeruleus and serotonergic raphe nuclei in S. salar. We did not find any coexistence of GALir substances with arginine vasotocin or isotocin in neurosecretory neurons, as has been shown for galanin with the mammalian counterparts vasopressin and oxytocin. Also, the galanin-like substance(s) and their structurally closest related peptide family, the tachykinins, belong to separate neuronal systems in teleosts. The presence of GALir neurons in brain areas known to be involved in pituitary control, and a massive GALir innervation of the pituitary, strongly indicate a role for galanin-like substances in pituitary control also in teleosts. Furthermore, the presence of extrahypothalamic GALir fibers suggests involvement of galanin-like substances in other brain functions in teleosts. In conclusion, there are general similarities between teleosts and mammals concerning the distribution of galanin-like substances. However, there seem to be substantial differences in their distribution relative to functionally related peptides within the hypothalamo-pituitary system. Whereas galanin appears to be colocalized and released together with vasopressin and oxytocin in mammals, in teleosts the homologous substances are contained within different sets of neurons that innervate the same target, the pituitary.

Clearance of the amyloid-beta peptide (A beta) as a remedy for Alzheimer's disease (AD) is a major target in on-going clinical trials. In vitro studies confirmed that A beta is taken up by rodent astrocytes, but knowledge on human astrocyte-mediated A beta clearance is sparse. Therefore, by means of flow cytometry and confocal laser scanning microscopy (CLSM), we evaluated the binding and internalization of A beta1-42 by primary human fetal astrocytes and adult astrocytes, isolated from nondemented subjects (n = 8) and AD subjects (n = 6). Furthermore, we analyzed whether alpha1-antichymotrypsin (ACT), which is found in amyloid plaques and can influence A beta fibrillogenesis, affects the A beta uptake by human astrocytes. Upon over night exposure of astrocytes to FAM-labeled A beta1-42 (10 microM) preparations, (80.7 +/- 17.7)% fetal and (52.9 +/- 20.9)% adult A beta-positive astrocytes (P = 0.018) were observed. No significant difference was found in A beta1-42 uptake between AD and non-AD astrocytes, and no influence of ApoE genotype on A beta1-42 uptake was observed in any group. There was no difference in the percentage of A beta-positive cells upon exposure to A beta1-42 (10 microM) combined with ACT (1,000:1, 100:1, and 10:1 molar ratio), versus A beta1-42 alone. CLSM revealed binding of A beta1-42 to the cellular surfaces and cellular internalization of smaller A beta1-42 fragments. Under these conditions, there was no increase in cellular release of the proinflammatory chemokine monocyte-chemoattractant protein 1, as compared with nontreated control astrocytes. Thus, primary human astrocytes derived from different sources can bind and internalize A beta1-42, and fetal astrocytes were more efficient in A beta1-42 uptake than adult astrocytes.

Results show that HAMLET is active as a tumoricidal agent and suggest that topical HAMLET administration may delay bladder cancer development.

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