Neurons take shape

To construct the intricate network of connections that supports the functions of an adult nervous system, neurons must form highly elaborate processes, extending in the appropriate direction across long distances to form synapses with their partners. As the nervous system takes shape, the process of neuronal morphogenesis is controlled by a broad repertoire of cellular signals. These extracellular cues and cellular interactions are translated by receptors at the cell surface into physical forces that control the dynamic architecture of the neuron as it explores the surrounding terrain. The interpretation of these cues involves a large set of intracellular proteins, whose functional logic we are just beginning to appreciate. We shall consider the basic mechanics of neuronal morphogenesis and some of the emerging pathways that seem to link the outer and inner worlds of the neuron.     

Respiration in Primary Cultured Cerebellar Granule Neurons and Cerebral Cortical Neurons

Respiration was measured polarographically in primary cultures enriched with cerebellar granule neurons or cerebral cortical neurons. The basal respiratory rate, measured on the sixth day after culturing, was 12.00 natom equiv. O/mg protein/min for the cortical neurons and 12.70 natom equiv. O/mg protein/min for the granule neurons. Maximal stimulation by 2,4-dinitrophenol produced a 20-40% increase over the basal rate for both neuronal types. Oligomycin inhibited neuronal basal respiration by 45%. These respiratory rates in neurons from primary culture are markedly lower than those measured in astrocytes grown under similar conditions.     

Postsynaptic Currents in Dorsal Root Ganglion Neurons Co-cultured with Spinal Cord Neurons

In co-cultured dorsal root ganglion (DRG) neurons and spinal cord neurons from newborn rats, using a voltage-clamp technique in the whole-cell configuration enabled us to observe in DRG neurons the effects evoked by extracellular local electrical stimulation of cells corresponding to spinal cord neurons in their morphological characteristics. Such stimulation caused the appearance of postsynaptic currents (PSC) in DRG neurons in 9% of the cases. The mean delay of these currents (measured from the stimulus leading edge) was 4.7 ± 0.29 msec, the mean time to peak was 2.6 ± 0.77 msec, and the decay time constant = 14.5 ± 1.04 msec. The reversal potential of evoked PSC (ePSC) was close to the equilibrium potential for chloride ions estimated by the Nernst equation. Application of 20 M bicuculline induced practically complete and reversible ePSC block. The conclusion was drawn that these currents arise due to activation of the chloride channels operated by GABA receptors and, hence, represent an inhibitory PSC. Thus, one may deem it proved that spinal cord neurons can establish functional inhibitory synapses with DRG neurons.     

Electrical Compartmentalization in Neurons

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The Reliability of Neurons

The prevalent probabilistic view is virtually untestable; it remains a plausible belief. The cases usually cited can not be taken as evidence for it. Several grounds for this conclusion are developed. Three issues are distinguished in an attempt to clarify a murky debate: (a) the utility of probabilistic methods in data reduction, (b) the value of models that assume indeterminacy, and (c) the validity of the inference that the nervous system is largely indeterministic at the neuronal level. No exception is taken to the first two; the second is a private heuristic question. The third is the issue to which the assertion in the first two sentences is addressed. Of the two kinds of uncertainty, statistical mechanical (= practical unpredictability) as in a gas, and Heisenbergian indeterminancy, the first certainly exists, the second is moot at the neuronal level. It would contribute to discussion to recognize that neurons perform with a degree of reliability. Although unreliability is difficult to establish, to say nothing of measure, evidence that some neurons have a high degree of reliability, in both connections and activity is increasing greatly. An example is given from sternarchine electric fish.     

Mouse Postganglionic Sympathetic Neurons

Basic properties of noradrenaline release were studied in primary cultures of thoracolumbar postganglionic sympathetic neurons taken from 1-3-day-old NMRI mice. After 7 days in vitro, the cultures were preincubated with [3H]noradrenaline and then superfused and stimulated electrically. Conventional trains of pulses (for example, 36 pulses at 3 Hz) as well as single pulses and brief high-frequency trains (for example, four pulses at 100 Hz) elicited a well-measurable overflow of tritium, which was abolished by 0.3 microM tetrodotoxin or omission of Ca2+, but not changed by 1 microM rauwolscine. In trains of one, two, four, six, eight, or 10 pulses at 3 Hz, the evoked overflow of tritium remained constant from pulse to pulse at 1.3 mM Ca2+, but declined slightly at 2.5 mM Ca2+. Tetraethylammonium at 10 mM selectively increased the overflow elicited by small pulse numbers and especially by a single pulse. In trains of 10 pulses delivered at 0.3, 1, 3, 10, 30, or 100 Hz, the evoked overflow of tritium increased from 0.3 to 30 Hz and then declined at 100 Hz. This relationship was particularly pronounced at low Ca2+ concentrations (for example, 0.3 mM). Tetraethylammonium at 10 mM selectively increased the overflow elicited by low frequencies of stimulation. It is concluded that primary cultures of mouse postganglionic sympathetic neurons can be used to investigate release of [3H]noradrenaline. The release is well measurable, even upon a single electrical pulse. It agrees with release in intact sympathetically innervated tissues in a number of fundamental properties, including the pulse number and frequency dependence. The preparation may be of special interest in conjunction with genetic manipulations in the donor animals.     

Mitochondrial Trafficking in Neurons

Neurons, perhaps more than any other cell type, depend on mitochondrial trafficking for their survival. Recent studies have elucidated a motor/adaptor complex on the mitochondrial surface that is shared between neurons and other animal cells. In addition to kinesin and dynein, this complex contains the proteins Miro (also called RhoT1/2) and milton (also called TRAK1/2) and is responsible for much, although not necessarily all, mitochondrial movement. Elucidation of the complex has permitted inroads for understanding how this movement is regulated by a variety of intracellular signals, although many mysteries remain. Regulating mitochondrial movement can match energy demand to energy supply throughout the extraordinary architecture of these cells and can control the clearance and replenishing of mitochondria in the periphery. Because the extended axons of neurons contain uniformly polarized microtubules, they have been useful for studying mitochondrial motility in conjunction with biochemical assays in many cell types.     

Spontaneous Activity in Crustacean Neurons

Single units which discharged with regular spontaneous rhythms without intentional stimulation were observed in the ventral nerve cord by intracellular recording close to the sixth abdominal ganglion. These units were divided into two groups: group A units in which interspike intervals varied less than 10 msec.; group B units in which interspike intervals varied within a range of 10 to 30 msec. Group A units maintained "constant" interspike intervals and could not be discharged by sensory inputs, while the majority of group B units could be discharged by appropriate sensory nerve stimulation. Both group A and B units discharged to direct stimulation when the stimulating and recording electrodes were placed in the same ganglionic intersegment, and directly evoked single spikes reset the spontaneous rhythm. In group B units, presynaptic volleys reset the spontaneous rhythm of some units; but in others, synaptically evoked spikes were interpolated within the spontaneous rhythm without resetting. The phenomenon of enhancement could also be demonstrated in spontaneously active units as a result of repetitive stimulation. It is concluded that endogenous pacemaker activity is responsible for much of the regular spontaneous firing observed in crayfish central neurons, and that interaction of evoked responses with such pacemaker sites can produce a variety of effects dependent upon the anatomical relationships between pacemaker and synaptic regions.     

Electrophysiology of Raccoon Cuneocerebellar Neurons

(1) Electrophysiological experiments were undertaken in order to locate and functionally characterize cells of the raccoon main cuneate nucleus (MCN) that can be activated by electrical stimulation of the cerebellum. A total of 98 such units were studied in pentobarbital sodium-anesthetized, methoxyflurane-anesthetized, or decerebrate preparations. Aside from a greater likelihood of resting discharge in the decerebrate preparations, no appreciable variability in physiological properties of the neurons could be attributed to differences in the type of preparation. (2) Using constant latency of response and ability to be blocked by collision as principal criteria, both antidromically (n = 31) and synaptically (n = 67) activated neurons of the main cuneate nucleus could be identified. A small number of MCN neurons could be activated by both cerebellar and thalamic stimulation, but no unit was antidromically activated from both locations. (3) MCN neurons projecting to the cerebellum are located primarily in the ventral polymorphic cell region of the nucleus at and rostral to the obex, corresponding to the “medial tongue” region of Johnson et al (1968). In contrast, neurons synaptically activated from the cerebellum are found throughout the dorsoventral extent of the rostral MCN, including the “clusters” region. (4) The majority of antidromically activated units responded to mechanical stimulation of deeper tissues, and most of these were activated by muscle stretch. Only a small portion (13-15%) of either antidromically or synaptically activated units were classed as light touch units with peripheral receptive fields (RFs) restricted to glabrous surfaces of the forepaw. (5) Glabrous skin RFs located on the digital surfaces are smaller than those located on the palm pads. In both cases, RFs are larger than those associated with primary afferent fibers, but toward the low end of the distribution for MCN neurons not activated by cerebellar stimulation. (6) All MCN units activated by cerebellar stimulation, regardless of modality, respond to mechanical stimulation with trains of irregularly spaced single spikes. Glabrous skin cutaneous mechanoreceptive MCN neurons, whether rapidly or slowly adapting, respond to ramp indentations with an instantaneous frequency which may be described as a power function of ramp velocity, with exponents less than one. These values are in the same range as those previously reported for primary afferents of the cuneate fasciculus (Pubols and Pubols, 1973).