Sonal Jhaveri

Whisker-related neuronal patterns fail to develop in the trigeminal brainstem nuclei of NMDAR1 knockout mice

Sensory pathways of the brain generally develop from crudely wired networks to precisely organized systems. Several studies have implicated neural activity-dependent mechanisms, including N-methyl-D-aspartate (NMDA) receptors, in this refinement process. We applied the gene targeting to the NMDAR1 gene and created a mutant mouse that lacks functional NMDA receptors. The development of whisker-related patterns in the trigeminal nuclei of the mutant mice and their normal littermates was compared. We show that in the mutant mice pathfinding, initial targeting, and crude topographic projection of trigeminal axons in the brainstem are unaffected, but that whisker-specific patches fail to form. Our results provide a direct demonstration of the involvement of the NMDA receptor in the formation of periphery-related neural patterns in the mammalian brain.

Thalamic axons confer a blueprint of the sensory periphery onto the developing rat somatosensory cortex

In order to study the role of afferents in the maturation of cortical axons projecting from the ventrobasal thalamic complex (VB) to the barrel field (SI) cortex were labeled with the carbocyanine dye DiI, in aldehyde-fixed embryonic and newborn rat brains. Our results reveal that the first few thalamic axons are in the cortical plate by embryonic day (E) 19. Between E19 and the day of birth (E21 = PND 0), layers V and VI differentiate from the lower part of the cortical plate. On PND 0, a plexus of growth-cone tipped thalamic axons is present within the cortical plate and a few VB fibers have reached the marginal zone. Increasing numbers of thalamic afferents invade and ramify within the cortical plate on PND 1 and, over the course of the next 24 h, form a vibrissa-specific pattern in the lower part of this zone, prior to the differentiation of layer IV into a distinct lamina. This periphery-related organization is exhibited by VB afferents earlier than reported for other afferents to the cortex, by glia- or neuron-associated extracellular elements or by the cytoarchitectonic specializations (barrels) of stellate cells. Our observations, in conjunction with the previous studies, demonstrate that thalamic afferents may have a pivotal role in determining the morphological specification of the primary somatosensory cortex.

Ablation of de novo DNA methyltransferase Dnmt3a in the nervous system leads to neuromuscular defects and shortened lifespan.

DNA methylation is an epigenetic mechanism involved in gene regulation and implicated in the functioning of the nervous system. The de novo DNA methyltransferase Dnmt3a is expressed in neurons, but its specific role has not been clarified. Dnmt3a is activated around embryonic day 10.5 in mouse neuronal precursor cells and remains active in postmitotic neurons in the adult. We assessed the role of neuronal Dnmt3a by conditional gene targeting. Mice lacking functional Dnmt3a in the nervous system were born healthy, but degenerated in adulthood and died prematurely. Mutant mice were hypoactive, walked abnormally, and underperformed on tests of neuromuscular function and motor coordination. Loss of Dnmt3a also led to fewer motor neurons in the hypoglossal nucleus and more fragmented endplates in neuromuscular junctions of the diaphragm muscle. Our results implicate a role for Dnmt3a in the neuromuscular control of motor movement. Developmental Dynamics 236:1663–1676, 2007.

Anomalous Retinal Pathways in the Siamese Cat: An Inadequate Substrate for Normal Binocular Vision

All major retinal pathways in the Siamese cat are abnormal, with almost total crossing of the projections to the pretectum and superior colliculus. These projections represent a marked disruption in the customary neural substrate for binocular vision, which implies a consequent impairment in stereoscopic depth perception. Crossed eyes, commonly seen in the Siamese cat, may therefore arise from a neuroanatomical defect in the primary visual pathways.

NOS inhibition during postnatal development leads to increased ipsilateral retinocollicular and retinogeniculate projections in rats

Synthesis of nitric oxide (NO) occurs downstream from activation of N‐methyl‐ d‐aspartate (NMDA) receptors; NO reportedly acts as a retrograde messenger, influencing the refinement and stabilization of coactive afferent terminals. Cells and neuropil in the rat superior colliculus (SC) and lateral geniculate body (LGB) show intense, developmentally regulated activity for NO synthase (NOS). To study the role of NO in the development of retinogeniculate and retinotectal axon arbors, we examined primary visual projections of rats that had received intraperitoneal injections of Nω‐nitro‐l‐arginine (L‐NoArg, an NOS inhibitor) on postnatal day 0, and daily thereafter for 4–6 weeks. Treated rats showed significant alterations in ipsilateral retinotectal projections, in the mediolateral and anteroposterior axes; there was an increase in the density of fibres entering the SC, in branch length, and in the numbers of boutons on retinotectal arbors in the treated group. Ipsilaterally projecting retinal axons also showed an increase in density and distribution in the dorsal nucleus of the LGB. If animals were allowed to survive for several months after stopping treatment, similar changes were also noted, but these were much less striking. Our results support the hypothesis that, in the mammalian visual system, NO released from target neurons in the SC and LGB serves as a retrograde signal which feeds back on retinal afferents, influencing their growth. The effects of NOS inhibition are partially reversed after treatment is stopped, indicating that lack of NO synthesis delays the maturation of retinofugal connections, and also that NO plays a constitutive role in their development.

Target-specific morphology of retinal axon arbors in the adult hamster

The B fragment of cholera toxin (CT-B) provides a highly sensitive anterograde tracer for labeling retinofugal axons, revealing dense projections to known central retinorecipient nuclei, and sparse but distinct inputs to regions that have not been traditionally recognized as targets of direct retinal projections. In hamsters, we can identify CT-B labeled retinal axons in more than 25 cell groups in the mesencephalon, diencephalon, and basal telencephalon. CT-B labeling additionally delineates the complete arbor morphology, especially in regions that receive a sparse input, offering hitherto unknown views of retinal axon ramifications. We present here the terminal morphology of retinal axons in the lateral geniculate body and superior colliculus, verifying earlier studies, and also document novel findings on the configuration of retinal axon endings in the ventral nucleus of the lateral geniculate body, intergeniculate leaflet, suprachiasmatic nucleus, and in the nuclei of the accessory optic tract. Additionally, the trajectory and terminal morphology of retinal afferents to the hypothalamus, preoptic area, and basal telencephalon are detailed. The results are discussed in the context of possible functional roles for some of these projections.

Afferents from the colliculus, cortex, and retina have distinct terminal morphologies in the lateral posterior thalamic nucleus

We have examined the morphology of afferent endings that originate in three distinct cell groups and terminate in the lateral posterior nucleus of the thalamus (LP). Retino‐LP projections were sparse, occurred throughout the nucleus, and could be classified into 1) simple en passant varicosities and terminal swellings found on poorly branched fibers in all LP subdivisions, 2) string‐like configurations of varicosities detected largely in the medial subdivision of the LP, and 3) terminals resembling retinogeniculate endings occurring mainly in the rostral part of the superficial subdivision of the LP adjacent to the dorsal nucleus of the lateral geniculate body. Cortico‐LP terminals fell into three classes: 1) single varicosities decorating the tips of short appendages on fine preterminal and terminal axons; 2) tiny, round varicosities studding the axon shaft; and 3) boutons of variable shape visible on medium‐caliber corticothalamic fibers. Tecto‐LP terminals exhibited a large variation in morphology and density. Those found most commonly could be classified into two groups: 1) individual swellings and 2) terminal clusters arranged in a tubular configuration that enclosed a central channel, most likely occupied by the dendrite of a postsynaptic neuron. An unusual tecto‐LP terminal consisted of an ovoid swelling (up to 20 μm in the long axis) from which emerged several long, thin extensions and was seen at the tips of large‐diameter axons. These results show that, despite having overlapping projection zones, each set of afferents that projects to the LP elaborates terminal specializations that are structurally distinct from others projecting to the same target area. J. Comp. Neurol. 388:467–483, 1997.

DEFECTIVE WHISKER FOLLICLES AND ALTERED BRAINSTEM PATTERNS IN ACTIVIN AND FOLLISTATIN KNOCKOUT MICE

Whisker pad innervation and whisker-specific pattern formation were examined in mice lacking the gene for activin betaA or for follistatin. Both strains of mice die within 24 h after birth. A normal array of whisker follicles is present in the snout of either phenotype. However, activin betaA-deficient mice lack whiskers, and in follistatin-deficient mice the whiskers are thin and curled. We examined the effects of aberrant, albeit innervated, follicles on the formation of whisker-specific patterns (barrelettes) in the trigeminal brainstem. Activin betaA knockout mice lack barrelettes, although the trigeminal afferent topography is not compromised. Physiological recordings suggest that trigeminal ganglion cells in these mice are less responsive to stimulation of whisker follicles. Barrelettes in follistatin-deficient mice are not as well developed as in controls, but can be discerned in some cases. These results are consistent with the notion that formation of barrelettes depends on neural activity initiated by the whiskers.

Target Influences on the Morphology of Trigeminal Axons

Axons grow in two stages: First, they exhibit rapid, target-directed extension; then they begin to collateralize and elaborate terminal arbors in their targets. To investigate possible regulatory influences on these phases of axon growth, we have used an in vitro paradigm in which we cocultured embryonic or postnatal rat trigeminal ganglion explants with isochronic, heterochronic, and/or heterotypic targets. Cultures were fixed after 5 days and ganglion cell processes were labeled with DiI. Trigeminal processes were able to regenerate into several peripheral targets as well as into CNS explants from trigeminal or nontrigeminal regions of the brain. In peripheral tissues, the processes showed target-specific growth patterns. In CNS tissue, the type of growth (unbranched extension versus collateralization/arbor formation) varied markedly with the explant age: If trigeminal ganglia were harvested at a time (E15) when their axons would be elongating in the embryo and cocultured with isochronic tissues, their processes had a simple morphology, were loosely bundled, and reconstituted a distinct fiber tract, mimicking their in vivo growth pattern. If, challenged by more mature tissue, axons of E15 ganglion cells formed discrete arbors. Finally, if trigeminal ganglia were harvested at an age (E20, PND 5) when their axons had already formed arbors in the brain and induced to innervate younger (E15) targets, their axons reverted back to the elongation stage. These results demonstrate that the target environment sets specific, developmentally regulated constraints on the patterns of growth manifested by primary sensory axons.

The gene knockout technology for the analysis of learning and memory, and neural development

Publisher Summary Using the gene knockout technology, this chapter presents an analysis of learning and memory, and neural development. It investigates the molecular substrates of synaptic plasticity by producing mice that lack the γ isoform of Ca 2+ /phospholipid-dependent-protein kinase (PKCγ) using the embryonic stem (ES) cell gene targeting technology. PKC constitutes a family of isoenzymes involved in signal transduction pathways in diverse systems. This enzyme was chosen for the study reviewed in the chapter because pharmacological studies have repeatedly implicated PKC as playing a role in long-term potentiation (LTP). To re-examine the role of N -methyl-D-aspartate (NMDA) receptor (NMDAR)-mediated activity in the establishment of neural patterns in the whisker-to-barrel system, reverse genetics is used to selectively “knock out” the NMDAR1 subunit of the NMDA receptor. The results from this examination show that in the knockout animals, although central targeting and topographic projection of trigeminal afferent appear to be normal and postsynaptic neurons are responsive to the stimulation of primary trigeminal afferent, whisker-specific neural patterns fail to develop in the absence of the NMDA receptor.