Activation of neurons in the hypothalamic dorsomedial nucleus via hypothalamic projections of the nucleus of the solitary tract following refeeding of fasted rats

We report that satiation evokes neuronal activity in the ventral subdivision of the hypothalamic dorsomedial nucleus (DMH) as indicated by increased c‐fos expression in response to refeeding in fasted rats. The absence of significant Fos activation following food presentation without consumption suggests that satiation but not craving for food elicits the activation of ventral DMH neurons. The distribution pattern of the prolactin‐releasing peptide (PrRP)‐immunoreactive (ir) network showed remarkable correlations with the distribution of activated neurons within the DMH. The PrRP‐ir fibers and terminals were immunolabeled with tyrosine hydroxylase, suggesting their origin in lower brainstem instead of local, hypothalamic PrRP cells. PrRP‐ir fibers arising from neurons of the nucleus of the solitary tract could be followed to the hypothalamus. Unilateral transections of these fibers at pontine and caudal hypothalamic levels resulted in a disappearance of the dense PrRP‐ir network in the ventral DMH while PrRP immunoreactivity was increased in transected fibers caudal to the knife cuts as well as in perikarya of the nucleus of the solitary tract ipsilateral to the transections. In accord with these changes, the number of Fos‐expressing neurons following refeeding declined in the ipsilateral but remained high in the contralateral DMH. However, the Fos response in the ventral DMH was not attenuated following chemical lesion (neonatal monosodium glutamate treatment) of the hypothalamic arcuate nucleus, another possible source of DMH inputs. These findings suggest that PrRP projections from the nucleus of the solitary tract contribute to the activation of ventral DMH neurons during refeeding, possibly by transferring information on cholecystokinin‐mediated satiation.     

Can neck muscle spindle afferents activate fusimotor neurons of the lower limb?

Introduction: We investigated whether vibratory stimulation of the dorsal neck muscles activates fusimotor neurons of lower limb muscles in relaxed human subjects. Methods: The triceps surae (TS) muscles of seated subjects (n = 15) were conditioned to leave their muscle spindles in either an insensitive (hold‐long) or sensitive (hold‐short) state. A vibrator (80 HZ ) was applied to the dorsal neck muscles for 10 seconds. The tendon jerk was evoked from the right TS immediately before (during) or 5 seconds after (interposed) the offset of vibration. Results: The size of the reflex after hold‐long muscle conditioning and after neck vibration was significantly smaller than the control hold‐short reflex (P < 0.001). However, after hold‐short conditioning, neck vibration significantly increased tendon jerk amplitude, both during (P = 0.001) and interposed (P = 0.026). Conclusion: Dorsal neck vibration increases spinal reflex excitability of the TS in relaxed and seated subjects, but not through fusimotor excitation. Muscle Nerve, 2012     

Paraplegia increases skeletal muscle autophagy

Introduction: Paraplegia results in significant skeletal muscle atrophy through increases in skeletal muscle protein breakdown. Recent work has identified a novel SIRT1–p53 pathway that is capable of regulating autophagy and protein breakdown. Methods: Soleus muscle was collected from 6 male Sprague‐Dawley rats 10 weeks after complete T4–5 spinal cord transection (paraplegia group) and 6 male sham‐operated rats (control group). We utilized immunoblotting methods to measure intracellular proteins and quantitative real‐time polymerase chain reaction to measure the expression of skeletal muscle microRNAs. Results: SIRT1 protein expression was 37% lower, and p53 acetylation (LYS379) was increased in the paraplegic rats (P < 0.05). Atg7 and Beclin‐1, markers of autophagy induction, were elevated in the paraplegia group compared with controls (P < 0.05). Conclusions: Severe muscle atrophy resulting from chronic paraplegia appears to increase skeletal muscle autophagy independent of SIRT1 signaling. We conclude that chronic paraplegia may cause an increase in autophagic cell death and negatively impact skeletal muscle protein balance. Muscle Nerve 46: 793–798, 2012     

Distribution of RET immunoreactivity in the rodent spinal cord and changes after nerve injury

RET (for “rearranged during transfection”) is a transmembrane tyrosine kinase signaling receptor for members of the glial cell line‐derived neurotrophic factor (GDNF) family of ligands. We used RET immunohistochemistry (IHC), double‐labeling immunofluorescence (IF), and in situ hybridization (ISH) in adult naïve and nerve‐injured rats to study the distribution of RET in the spinal cord. In the dorsal horn, strong RET‐immunoreactive (‐ir) fibers were abundant in lamina II‐inner (II_i), although this labeling was preferentially observed after an antigen‐unmasking procedure. After dorsal rhizotomy, RET‐ir fibers in lamina II_i completely disappeared from the dorsal horn, indicating that they were all primary afferents. After peripheral axotomy, RET‐ir in primary afferents decreased in lamina II_i and appeared to increase slightly in laminae III and IV. RET‐ir was also observed in neurons and dendrites throughout the dorsal horn. Some RET‐ir neurons in lamina I had the morphological appearance of nociceptive projection neurons, which was confirmed by the finding that 53% of RET‐ir neurons in lamina I colocalized with neurokinin‐1. GDNF‐ir terminals were in close proximity to RET‐ir neurons in the superficial dorsal horn. In the ventral horn, RET‐ir was strongly expressed by motoneurons, with the strongest staining in small, presumably γ‐motoneurons. Increased RET expression following peripheral axotomy was most pronounced in α‐motoneurons. The expression and regulation pattern of RET in the spinal cord are in line with its involvement in regenerative processes following nerve injury. The presence of RET in dorsal horn neurons, including nociceptive projection neurons, suggests that RET also has a role in signal transduction at the spinal level. This role may include mediating the effects of GDNF released from nociceptive afferent fibers. J. Comp. Neurol. 500:1136–1153, 2007. © 2006 Wiley‐Liss, Inc.     

Activation of the Locus Coeruleus After Amygdaloid Kindling

Summary: Purpose: Substnatia nigra (SN) and locus coeruleus (LC) neurons are implicated in the propagation and suppression of amygdaloid seizures. Both structures are activated concomitant with amygdaloid seizure discharges. Their rnechanisms of activation, however, remain to be elucidated. SN firing is not associated with the induction of Fos imrnunoreactiv-ity (ir), a marker of excitatory neuronal activation. LC has not been studied. The purpose of this investigation was to determine if amygdala-kindled generalized seizures could induce Fos-ir in the LC. Methods: Female Sprague-Dawley rats were killed after generalized seizures induced by amygdala electrical stimulation and stained by using Fos irnmunocytochemistry. The number of Fos-ir neurons was compared between 15 animals with generalized seizures and four implanted, unstimulated controls. Results: LC-ir neurons were significantly (p < 0.05) more prevalent after seizures than in control animals. Their numbers correlated very highly with Fos-ir in the central nucleus of the amygdala (p < 0.0001). No Fos induction was observed in LC in controls or in the SN in either group. Conclusions: Amygdala-induced generalized seizures result in Fos-ir in the LC but not in the SN. This is consistent with different mechanisms of activation possibly involving disinhi-bition in the SN and direct excitation in the LC.     

Different patterns of neuronal activation and neurodegeneration in the thalamus and cortex of epilepsy‐resistant Proechimys rats versus Wistar rats after pilocarpine‐induced protracted seizures

Purpose: To analyze cellular mechanisms of limbic‐seizure suppression, the response to pilocarpine‐induced seizures was investigated in cortex and thalamus, comparing epilepsy‐resistant rats Proechimys guyannensis with Wistar rats. Methods: Fos immunoreactivity revealing neuronal activation, and degenerating neurons labeled by Fluoro‐Jade B (FJB) histochemistry were analyzed on the first day after onset of seizures lasting 3 h. Subpopulations of γ‐aminobutyric acid (GABA)ergic cells were characterized with double Fos‐parvalbumin immunohistochemistry. Results: In both cortex and thalamus, degenerating neurons were much fewer in Proechimys than Wistar rats. Fos persisted at high levels at 24 h only in the Proechimys thalamus and cortex, especially in layer VI where corticothalamic neurons reside. In the parietal cortex, about 50% of parvalbumin‐containing interneurons at 8 h, and 10–20% at 24 h, were Fos‐positive in Wistar rats, but in Proechimys, Fos was expressed in almost all parvalbumin‐containing interneurons at 8 h and dropped at 24 h. Fos positivity in cingulate cortex interneurons was similar in both species. In the Wistar rat thalamus, Fos was induced in medial and midline nuclei up to 8 h, when <30% of reticular nucleus cells were Fos‐positive, and then decreased, with no relationship with cell loss, evaluated in Nissl‐stained sections. In Proechimys, almost all reticular nucleus neurons were Fos‐positive at 24 h. Discussion: At variance with laboratory rats, pilocarpine‐induced protracted seizures elicit in Proechimys limited neuronal death, and marked and long‐lasting Fos induction in excitatory and inhibitory cortical and thalamic cell subsets. The findings implicate intrathalamic and intracortical regulation, and circuits linking thalamus and cortex in limbic seizure suppression leading to epilepsy resistance.     

Comparative mapping of GABA‐immunoreactive neurons in the central nervous systems of nudibranch molluscs

The relative simplicity of certain invertebrate nervous systems, such as those of gastropod molluscs, allows behaviors to be dissected at the level of small neural circuits composed of individually identifiable neurons. Elucidating the neurotransmitter phenotype of neurons in neural circuits is important for understanding how those neural circuits function. In this study, we examined the distribution of γ‐aminobutyric‐acid;‐immunoreactive (GABA‐ir) neurons in four species of sea slugs (Mollusca, Gastropoda, Opisthobranchia, Nudibranchia): Tritonia diomedea, Melibe leonina, Dendronotus iris, and Hermissenda crassicornis. We found consistent patterns of GABA immunoreactivity in the pedal and cerebral‐pleural ganglia across species. In particular, there were bilateral clusters in the lateral and medial regions of the dorsal surface of the cerebral ganglia as well as a cluster on the ventral surface of the pedal ganglia. There were also individual GABA‐ir neurons that were recognizable across species. The invariant presence of these individual neurons and clusters suggests that they are homologous, although there were interspecies differences in the numbers of neurons in the clusters. The GABAergic system was largely restricted to the central nervous system, with the majority of axons confined to ganglionic connectives and commissures, suggesting a central, integrative role for GABA. GABA was a candidate inhibitory neurotransmitter for neurons in central pattern generator (CPG) circuits underlying swimming behaviors in these species, however none of the known swim CPG neurons were GABA‐ir. Although the functions of these GABA‐ir neurons are not known, it is clear that their presence has been strongly conserved across nudibranchs. J. Comp. Neurol. 522:794–810, 2014. © 2013 Wiley Periodicals, Inc.     

Comparative Mapping of GABA‐Immunoreactive Neurons in the Buccal Ganglia of Nudipleura Molluscs

Phylogenetic comparisons of neurotransmitter distribution are important for understanding the ground plan organization of nervous systems. This study describes the γ‐aminobutyric acid (GABA)‐immunoreactive (GABA‐ir) neurons in the buccal ganglia of six sea slug species (Mollusca, Gastropoda, Euthyneura, Nudipleura). In the nudibranch species, Hermissenda crassicornis, Tritonia diomedea, Tochuina tetraquetra, and Dendronotus iris, the number of GABA‐ir neurons was highly consistent. Another nudibranch, Melibe leonina, however, contained approximately half the number of GABA‐ir neurons. This may relate to its loss of a radula and its unique feeding behavior. The GABA immunoreactivity in a sister group to the nudibranchs, Pleurobranchaea californica, differed drastically from that of the nudibranchs. Not only did it have significantly more GABA‐ir neurons but it also had a unique GABA distribution pattern. Furthermore, unlike the nudibranchs, the Pleurobranchaea GABA distribution was also different from that of other, more distantly related, euopisthobranch and panpulmonate snails and slugs. This suggests that the Pleurobranchaea GABA distribution may be a derived feature, unique to this lineage. The majority of GABA‐ir axons and neuropil in the Nudipleura were restricted to the buccal ganglia, commissures, and connectives. However, in Tritonia and Pleurobranchaea, we detected a few GABA‐ir fibers in buccal nerves that innervate feeding muscles. Although the specific functions of the GABA‐ir neurons in the species in this study are not known, the innervation pattern suggests these neurons may play an integrative or regulatory role in bilaterally coordinated behaviors in the Nudipleura. J. Comp. Neurol. 524:1181–1192, 2016. © 2015 Wiley Periodicals, Inc.     

Developmental maturation of ionotropic glutamate receptor subunits in rat vestibular nuclear neurons responsive to vertical linear acceleration

We investigated the maturation profile of subunits of ionotropic glutamate receptors in vestibular nuclear neurons that were activated by sinusoidal linear acceleration along the vertical plane. The otolithic origin of Fos expression in these neurons was confirmed as a marker of functional activation when labyrinthectomized and/or stationary control rats contrasted by showing sporadically scattered Fos‐labeled neurons in the vestibular nuclei. By double immunohistochemistry for Fos and one of the receptor subunits, otolith‐related neurons that expressed either α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole‐propionate or N‐methyl‐d ‐aspartate subunits were first identified in the medial vestibular nucleus, spinal vestibular nucleus and Group x by postnatal day (P)7, and in the lateral vestibular nucleus and Group y by P9. No double‐labeled neurons were found in the superior vestibular nucleus. Within each vestibular subnucleus, these double‐labeled neurons constituted ～90% of the total Fos‐labeled neurons. The percentage of Fos‐labeled neurons expressing the GluR1 or NR2A subunit showed developmental invariance in all subnuclei. For Fos‐labeled neurons expressing the NR1 subunit, similar invariance was observed except that, in Group y, these neurons decreased from P14 onwards. For Fos‐labeled neurons expressing the GluR2, GluR2/3, GluR4 or NR2B subunit, a significant decrease was found by the adult stage. In particular, those expressing the GluR4 subunit showed a two‐ to threefold decrease in the medial vestibular nucleus, spinal vestibular nucleus and Group y. Also, those expressing the NR2B subunit showed a twofold decrease in Group y. Taken together, the postsynaptic expression of ionotropic glutamate receptor subunits in different vestibular subnuclei suggests that glutamatergic transmission within subregions plays differential developmental roles in the coding of gravity‐related vertical spatial information.     

Organization of alpha‐transducin immunoreactive system in the brain and retina of larval and young adult Sea Lamprey (Petromyzon marinus), and their relationship with other neural systems

We employed an anti‐transducin antibody (Gαt‐S), in combination with other markers, to characterize the Gαt‐S‐immunoreactive (ir) system in the CNS of the sea lamprey, Petromyzon marinus. Gαt‐S immunoreactivity was observed in some neuronal populations and numerous fibers distributed throughout the brain. Double Gαt‐S‐ and opsin‐ir neurons (putative photoreceptors) are distributed in the hypothalamus (postoptic commissure nucleus, dorsal and ventral hypothalamus) and caudal diencephalon, confirming results of García‐Fernández et al. (Cell and Tissue Research, 288, 267–278, 1997). Singly Gαt‐S‐ir cells were observed in the midbrain and hindbrain, increasing the known populations. Our results reveal for the first time in vertebrates the extensive innervation of many brain regions and the spinal cord by Gαt‐S‐ir fibers. The Gαt‐S innervation of the habenula is very selective, fibers densely innervating the lamprey homologue of the mammalian medial nucleus (Stephenson‐Jones et al., Proceedings of the National Academy of Sciences of the United States of America, 109, E164–E173, 2012), but not the lateral nucleus homologue. The lamprey neurohypophysis was not innervated by Gαt‐S‐ir fibers. We also analyzed by double immunofluorescence the relation of this system with other systems. A dopaminergic marker (TH), serotonin (5‐HT) or GABA do not co‐localize with Gαt‐S‐ir neurons although codistribution of fibers was observed. Codistribution of Gαt‐S‐ir fibers and isolectin‐labeled extrabulbar primary olfactory fibers was observed in the striatum and hypothalamus. Neurobiotin retrograde transport from the spinal cord combined with immunofluorescence revealed spinal‐projecting Gαt‐S‐ir reticular neurons in the caudal hindbrain. Present results in an ancient vertebrate reveal for the first time a collection of brain targets of Gαt‐S‐ir neurons, suggesting they might mediate non‐visual modulation by light in many systems.     

Wake‐promoting actions of noradrenergic α1‐ and β‐receptors within the lateral hypothalamic area

Central norepinephrine exerts potent wake‐promoting effects, in part through the actions of noradrenergic α₁‐ and β‐receptors located in the medial septal and medial preoptic areas. The lateral hypothalamic area (LHA), including the lateral hypothalamus, perifornical area and adjacent dorsomedial hypothalamus, is implicated in the regulation of arousal and receives a substantial noradrenergic innervation. To date the functional significance of this innervation is unknown. The current studies examined the degree to which noradrenergic α₁‐ and β‐receptor stimulation within the rat LHA modulates arousal. Specifically, these studies examined the wake‐promoting effects of intra‐tissue infusions (250 nL) of the α₁‐receptor agonist phenylephrine (10, 20 and 40 nmol) and the β‐receptor agonist isoproterenol (3, 10 and 30 nmol) in rats. Results show that stimulation of LHA α₁‐receptors elicits robust and dose‐dependent increases in waking. In contrast, β‐receptor stimulation within the LHA had relatively modest arousal‐promoting actions. Nonetheless, combined α₁‐ and β‐receptor stimulation elicited additive wake‐promoting effects. Arousal‐promoting hypocretin/orexin (HCRT)‐synthesising neurons are located within the LHA. Therefore, additional immunohistochemical studies examined whether α₁‐receptor‐dependent waking is associated with an activation of HCRT neurons as measured by Fos , the protein product of the immediate–early gene c‐fos. Analyses indicate that although intra‐LHA α₁‐receptor agonist infusion elicited a robust increase in Fos immunoreactivity (ir) in this region, this treatment did not activate HCRT neurons as measured by Fos‐ir. Collectively, these observations indicate that noradrenergic α₁‐receptors within the LHA promote arousal via actions that are independent of HCRT neuronal activation.     

7‐T MRI of the spinal cord can detect lateral corticospinal tract abnormality in amyotrophic lateral sclerosis

Introduction: Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder affecting lower and upper motor neurons. Degeneration of the lateral corticospinal tract (CST) is a key finding in ALS cervical spinal cord autopsies. We hypothesized that in vivo ultra‐high‐field MRI of the cervical spinal cord can detect abnormality in the CST. Methods: A patient with ALS (disease duration 23 months) and a healthy control were scanned at 7‐T MRI using a 19‐channel coil. Multi‐echo ‐weighted imaging was performed in the spinal cord, covering C2–C6. Cross‐sectional resolution was 0.37 × 0.37 mm². Results: We detected clear signal hyperintensity in both segments of the lateral CST in the ALS patient, which was significant when compared with the normal control subject (P < 10^?7). Conclusion: We believe there are potential benefits of 7‐T MRI for increased sensitivity and spatial accuracy in characterizing pathology in the spinal cord. Muscle Nerve 47: 760–762, 2013     

Degeneration of proprioceptive sensory nerve endings in mice harboring amyotrophic lateral sclerosis–causing mutations

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that primarily targets the motor system. Although much is known about the effects of ALS on motor neurons and glial cells, little is known about its effect on proprioceptive sensory neurons. This study examines proprioceptive sensory neurons in mice harboring mutations associated with ALS, in SOD1^G93A and TDP43^A315T transgenic mice. In both transgenic lines, we found fewer proprioceptive sensory neurons containing fluorescently tagged cholera toxin in their soma five days after injecting this retrograde tracer into the tibialis anterior muscle. We asked whether this is due to neuronal loss or selective degeneration of peripheral nerve endings. We found no difference in the total number and size of proprioceptive sensory neuron soma between symptomatic SOD1^G93A and control mice. However, analysis of proprioceptive nerve endings in muscles revealed early and significant alterations at Ia/II proprioceptive nerve endings in muscle spindles before the symptomatic phase of the disease. Although these changes occur alongside those at α‐motor axons in SOD1^G93A mice, Ia/II sensory nerve endings degenerate in the absence of obvious alterations in α‐motor axons in TDP43^A315T transgenic mice. We next asked whether proprioceptive nerve endings are similarly affected in the spinal cord and found that nerve endings terminating on α‐motor neurons are affected during the symptomatic phase and after peripheral nerve endings begin to degenerate. Overall, we show that Ia/II proprioceptive sensory neurons are affected by ALS‐causing mutations, with pathological changes starting at their peripheral nerve endings. J. Comp. Neurol. 523:2477–2494, 2015. © 2015 Wiley Periodicals, Inc.     

Soleus and lateral gastrocnemius H‐reflexes during standing with unstable footwear

Introduction: Unstable footwear has been shown to increase lower extremity muscle activity, but the reflex response to perturbations induced by this intervention is unknown. Methods: Twenty healthy subjects stood in stable and unstable footwear conditions (presented randomly) while H‐reflex amplitude and background muscle activity were measured in the soleus and lateral gastrocnemius (LG) muscles. Results: Wearing unstable footwear resulted in larger H‐reflexes (normalized to the maximal M‐wave) for the LG (+12%; P = 0.025), but not for the soleus (+4%; P > 0.05). Background activity of both muscles was significantly higher in the unstable condition. Conclusions: The H‐reflex facilitation observed with unstable footwear was unexpected, as challenging postural conditions usually result in reflex depression. Increased muscle activity, decreased presynaptic inhibition, and/or more forward postural position may have (over‐)compensated the expected reflex depression. Differences between LG and soleus H‐reflex modulation may be due to diverging motor unit recruitment thresholds. Muscle Nerve 51 :764–766, 2015     

Changes in GABA and GABA(B) receptor expressions are involved in neuropathy in the rat cuneate nucleus following median nerve transection

This study examined the relationship between changes in GABA transmission and behavioral abnormalities after median nerve transection. Following unilateral median nerve transection, the percentage of GABA‐like immunoreactive neurons in the cuneate nucleus and that of GABA_B receptor‐like immunoreactive neurons in the dorsal root ganglion in the injured side decreased and reached a nadir at 4 weeks after median nerve transection. Four weeks after bilateral median nerve transection and intraperitoneal application with saline, baclofen (2 mg kg^?1), or phaclofen (2 mg kg^?1) before unilateral electrical stimulation of the injured median nerve, we investigated the level of neuropeptide Y release and c‐Fos expression in the stimulated side of the cuneate nucleus. The neuropeptide Y release level and the number of c‐Fos‐like immunoreactive neurons in the baclofen group were significantly attenuated, whereas those in the phaclofen group had increased compared to the saline group. These findings indicate that median nerve transection reduces GABA transmission, promoting injury‐induced neuropeptide Y release and consequently evoking c‐Fos expression in cuneate nucleus neurons. Furthermore, this study used the CatWalk method to assess behavioral abnormalities in rats following median nerve transection. These abnormalities were reversed by baclofen treatment. Overall, the results suggest that baclofen treatment block neuropeptide Y release, subsequently lessening c‐Fos expression in cuneate neurons and consequently attenuating neuropathic signal transmission to the thalamus. Synapse, 2012. © 2012 Wiley Periodicals, Inc.     

Assessment of musculo‐articular and muscle stiffness in young and older men

Introduction: The aim of this cross‐sectional study was to concurrently assess musculo‐articular stiffness (MAS) and muscle stiffness (MS) of the knee extensors in younger and older individuals. Methods: Fourteen young (22.1 ± 3.0 years old) and 12 older (65.4 ± 5.7 years old) men were tested for maximal voluntary contraction (MVC), rate of torque development (RTD), muscle thickness, MAS, and MS of knee extensors. Results: MVC, RTD, and muscle thickness were higher in the younger group (288.6 vs. 194.3 Nm, 1319.5 vs. 787.0 Nm s^?1, 23.1 vs. 17.7 mm, respectively, P < 0.05). MAS normalized to the load supported (30% of MVC) was not different between groups (87.9 vs. 88.5 Nm^?1kg^?1), whereas the older group exhibited a higher level of normalized MS (23.2 vs. 18.6 Nm^?1kg^?1, P < 0.05). Conclusions: Determinants of MS have been highlighted along with their role in elevated MS. The unaltered level of MAS, which is functionally important in an aging population, might be achieved through a decrease in tendon stiffness. Muscle Nerve 46: 559–565, 2012