Formation of neurites and their differentiation into axons and dendrites requires precisely controlled changes in the cytoskeleton. While small GTPases of the Rho family appear to be involved in this regulation, it is still unclear how Rho function affects axonal and dendritic growth during development. Using hippocampal neurones at defined states of differentiation, we have dissected the function of RhoA in axonal and dendritic growth. Expression of a dominant negative RhoA variant inhibited axonal growth, whereas dendritic growth was promoted. The opposite phenotype was observed when a constitutively active RhoA variant was expressed. Inactivation of Rho by C3-catalysed ADP-ribosylation using C3 isoforms (Clostridium limosum, C3 lim orStaphylococcus aureus, C3 stau2 ), diminished axonal branching. By contrast, extracellularly applied nanomolar concentrations of C3 from C. botulinum (C3 bot ) or enzymatically dead C3 bot significantly increased axon growth and axon branching. Taken together, axonal development requires activation of RhoA, whereas dendritic development benefits from its inactivation. However, extracellular application of enzymatically active or dead C3 bot exclusively promotes axonal growth and branching suggesting a novel neurotrophic function of C3 that is independent from its enzymatic activity.

Changes in extracellular potassium concentration as measured with ion-selective microelectrodes revealed abnormally large accumulations in the hippocampus during postnatal development. While rises in [K' I, during stimulation of the Schaffer collaterals were limited to about 12 mM in adult animals, identical stimulations elicited rises to levels as large as 18 mM in juveniles. Since astrocytes are believed to play an important role in K+ homeostasis, we studied the postnatal development of astrocytes in the CA1 region of rat hippocampus in four age groups using a polyclonal antibody against glial fibrillary acidic protein (GFAP). The main proliferation of GFAP-positive cells (GFAPpc) occurred in all laminae between postnatal days 8 and 16. The numer of GFAP-positive astrocytes per unit area was reached in stratum lacunosum-moleculare and stratum oriens at about 2 weeks and in stratum radiatum at about 3 weeks of age. During further development-at the age of 24 days-the orientation of individual astrocytes in stratum radiatum became polar with an orientation almost perpendicular to stratum pyramidale. This was revealed by an analysis based on determination of the quotients between the angular orientation of the processes of single individual GFAPpositive cells. When the crossing points of all glial processes over vertical and horizontal grid lines were determined and respective quotients evaluated, the same development towards a perpendicular orientation of astrocytes was noted in stratum radiatum. The same approach revealed a transient orientation parallel to the fissure in stratum lacunosum-moleculare around day 24. Camera lucida drawings of GFAPpc in stratum radiatum revealed that astrocytes became larger during the first three postnatal weeks, followed by a reduction of various parameters (e.g., cell extension, branching pattern) until adulthood. The observed developmental changes of astroglial cells may contribute to the known delayed maturation of potassium regulation in rat hippocampus.

Non-technical summary Deep brain stimulation (DBS) refers to a neurosurgical technique where chronically implanted electrodes serve to deliver electrical impulses to highly defined brain regions in neuropsychiatric disorders including Parkinson's disease (PD), depression and obsessive-compulsive disorders. Despite its broad acceptance as a safe and effective treatment for advanced PD, DBS has remained enigmatic with respect to its underlying mechanism(s). In general terms, DBS is capable of reinstating regular electrical activity in the complex neuronal networks that exhibit aberrant firing properties in PD, but how this effect is achieved at the network and cellular level is still highly debated. Our study focuses on a crucial, but hitherto neglected, issue in this controversy, namely the propagation of impulses within and away from the site of electrical stimulation. We propose that DBS overburdens the capacity of axons to transmit signals, thereby filtering and abating the pathological activity in the brain motor loops of PD patients.Abstract Deep brain stimulation (DBS) has been established as an effective surgical therapy for advanced Parkinson's disease (PD) and gains increasing acceptance for otherwise intractable neuropsychiatric diseases such as major depression or obsessive-compulsive disorders. In PD, DBS targets predominantly the subthalamic nucleus (STN) and relieves motor deficits only at high frequency (>100 Hz). In contrast to the well-documented clinical efficacy of DBS, its underlying principle remains enigmatic spawning a broad and, in part, contradictory spectrum of suggested synaptic and non-synaptic mechanisms within and outside STN. Here we focused on a crucial, but largely neglected issue in this controversy, namely the axonal propagation of DBS within and away from STN. In rat brain slices preserving STN projections to substantia nigra (SN) and entopeduncular nucleus (EP, the rodent equivalent of internal globus pallidus), STN-DBS disrupted synaptic excitation onto target neurons through an unexpected failure of axonal signalling. The rapid onset and, upon termination of DBS, recovery of this effect was highly reminiscent of the time course of DBS in the clinical setting. We propose that DBS-induced suppression of axonal projections from and to STN serves to shield basal ganglia circuitry from pathological activity arising in or amplified by this nucleus.

Song control regions in passerine birds are known to be sexually dimorphic in the adult brain in species like zebra finches in which most males sing whereas females do not. As a first step toward the analysis of the establishment of anatomical sex differences, volumetric changes of Nissl-defined song control regions in the forebrain of the zebra finch have been quantified in both sexes at 10-day intervals starting at day 10 posthatching. In males, the Nissl-defined volume of the high vocal center, the robust nucleus of the archistriatum, and area X of the lobus parolfactorius increased with age, reaching the adult value at 60, 50, and at 40 days posthatching, respectively. The lateral magnocellular nucleus of the anterior neostriatum increased in volume until 20 days and then gradually decreased to reach the adult value at about 40 days. Whereas area X is absent in females, the high vocal center, the robust nucleus of the archistriatum, and the lateral magnocellular nucleus of the anterior neostriatum were detectable throughout development and in adulthood. In contrast to the males, volumes of the high vocal center and of the robust nucleus of the archistriatum decreased in females between 10 and 40 days posthatching (58% and 61%, respectively), when adult values were reached. Contrary to the development of these two nuclei in females, the volumetric development of the lateral magnocellular nucleus of the anterior neostriatum was very similar in both sexes. Females began with a smaller lateral magnocellular nucleus of the anterior neostriatum at 10 days posthatch, which led to a sexual dimorphism in juvenile stages, but there was no sexual dimorphism of volume size in adult brains.

We investigated the development of spiny neurons in the lateral magnocellular nucleus of the anterior neostriatum before, during, and after song learning in male zebra finches (Taeniopygia guttata). The frequency of dendritic spines, dendritic field size, and branching characteristics were quantified at different ages in Golgi-stained tissue using a three-dimensional computerized tracing system. During development, overall spine frequencies increase between 3 and 5 weeks and decrease thereafter. In particular, spine frequencies of middle segments decrease significantly by 14% between 5 and 7 weeks posthatching (p = 0.017). A further reduction of 48% occurs between 7 weeks and adulthood (p < 0.001), resulting in a spine reduction of 56% on middle segments between 35 days of age and adulthood. In addition to the reduction of spine frequencies, we find regressive events also on some of the neuronal parameters that we have quantified. In general, dendrites of adult animals terminate closer to the cell body than those of 7-, 5-, or 3-week-old birds. Whereas no changes in segment length of first- and second-order dendrites have been identified, third-order dendrites end 19% closer to the cell body in adults than in younger birds (p < 0.024). Second-order dendrites in adult animals branch less frequently than in 3-week-old animals (35%, p = 0.017). There is also a trend of a smaller number of tertiary branches in adulthood compared with 3-week-old birds (41%, p = 0.060). The morphological changes may be related to the function of this nucleus and the sensitive phase for song acquisition.

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BackgroundCGRP is contained in a substantial proportion of unmyelinated trigeminal neurons innervating intracranial tissues. Previously, we have described a hemisected rodent scull preparation and later the intact trigeminal ganglion to measure stimulated CGRP release from trigeminal afferents.MethodsHere, we establish a preparation for examining CGRP release from central trigeminal terminals using single fresh slices of the mouse medullary brainstem.ResultsBasal and stimulated amount of CGRP substantially exceeded the detection level. Experiments were designed as matched pairs of at least six brainstem slices per animal. Stimulation with high potassium induced calcium-dependent and reversible CGRP release. Capsaicin stimulation of TRPV1 provoked concentration-dependent CGRP release. The anti-migraine drug naratriptan did not inhibit capsaicin-induced CGRP release from peripheral terminals but inhibited the release from brainstem slices. The glutamate antagonist kynurenate showed a similar pattern of site-specific inhibition of CGRP release.ConclusionsAs observed earlier for other drugs used in the treatment of migraine this indicates that the central terminals in the spinal trigeminal nucleus may be the main site of action. The preparation allows evaluating the trigeminal brainstem as a pharmacological site of action.

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