In Vivo Calcium Imaging Visualizes Incision-Induced Primary Afferent Sensitization and Its Amelioration by Capsaicin Pretreatment.

Previous studies have shown that infiltration of capsaicin into the surgical site can prevent incision-induced spontaneous pain like behaviors and heat hyperalgesia. In the present study, we aimed to monitor primary sensory neuron Ca2+ activity in the intact dorsal root ganglia (DRG) using Pirt-GCaMP3 male and female mice pretreated with capsaicin or vehicle before the plantar incision. Intraplantar injection of capsaicin (0.05%) significantly attenuated spontaneous pain, mechanical, and heat hypersensitivity after plantar incision. The Ca2+ response in in vivo DRG and in in situ spinal cord was significantly enhanced in the ipsilateral side compared with contralateral side or naive control. Primary sensory nerve fiber length was significantly decreased in the incision skin area in capsaicin-pretreated animals detected by immunohistochemistry and placental alkaline phosphatase (PLAP) staining. Thus, capsaicin pretreatment attenuates incisional pain by suppressing Ca2+ response because of degeneration of primary sensory nerve fibers in the skin.SIGNIFICANCE STATEMENT Postoperative surgery pain is a major health and economic problem worldwide with ∼235 million major surgical procedures annually. Approximately 50% of these patients report uncontrolled or poorly controlled postoperative pain. However, mechanistic studies of postoperative surgery pain in primary sensory neurons have been limited to in vitro models or small numbers of neurons. Using an innovative, distinctive, and interdisciplinary in vivo populational dorsal root ganglia (DRG) imaging (>1800 neurons/DRG) approach, we revealed increased DRG neuronal Ca2+ activity from postoperative pain mouse model. This indicates widespread DRG primary sensory neuron plasticity. Increased neuronal Ca2+ activity occurs among various sizes of neurons but mostly in small-diameter and medium-diameter nociceptors. Capsaicin pretreatment as a therapeutic option significantly attenuates Ca2+ activity and postoperative pain.

Dynamics of the fragile X mental retardation protein correlates with cellular and synaptic properties in primary auditory neurons following afferent deprivation.

Afferent activity dynamically regulates neuronal properties and connectivity in the central nervous system. The Fragile X mental retardation protein (FMRP) is an RNA-binding protein that regulates cellular and synaptic properties in an activity-dependent manner. Whether and how FMRP level and localization are regulated by afferent input remains sparsely examined and how such regulation is associated with neuronal response to changes in sensory input is unknown. We characterized changes in FMRP level and localization in the chicken nucleus magnocellularis (NM), a primary cochlear nucleus, following afferent deprivation by unilateral cochlea removal. We observed rapid (within 2 hr) aggregation of FMRP immunoreactivity into large granular structures in a subset of deafferented NM neurons. Neurons that exhibited persistent FMRP aggregation at 12-24 hr eventually lost cytoplasmic Nissl substance, indicating cell death. A week later, FMRP expression in surviving neurons regained its homeostasis, with a slightly reduced immunostaining intensity and enhanced heterogeneity. Correlation analyses under the homeostatic status (7-14 days) revealed that neurons expressing relatively more FMRP had a higher capability of maintaining cell body size and ribosomal activity, as well as a better ability to detach inactive presynaptic terminals. Additionally, the intensity of an inhibitory postsynaptic protein, gephyrin, was reduced following deafferentation and was positively correlated with FMRP intensity, implicating an involvement of FMRP in synaptic dynamics in response to reduced afferent inputs. Collectively, this study demonstrates that afferent input regulates FMRP expression and localization in ways associated with multiple types of neuronal responses and synaptic rearrangements.

Mapping of the excitatory, inhibitory, and modulatory afferent projections to the anatomically defined active expiratory oscillator in adult male rats.

The lateral parafacial region (pFL ; which encompasses the parafacial respiratory group, pFRG) is a conditional oscillator that drives active expiration during periods of high respiratory demand, and increases ventilation through the recruitment of expiratory muscles. The pFL activity is highly modulated, and systematic analysis of its afferent projections is required to understand its connectivity and modulatory control. We combined a viral retrograde tracing approach to map direct brainstem projections to the putative location of pFL , with RNAScope and immunofluorescence to identify the neurochemical phenotype of the projecting neurons. Within the medulla, retrogradely-labeled, glutamatergic, glycinergic and GABAergic neurons were found in the ventral respiratory column (Bötzinger and preBötzinger Complex [preBötC], ventral respiratory group, ventral parafacial region [pFV ] and pFL ), nucleus of the solitary tract (NTS), reticular formation (RF), pontine and midbrain vestibular nuclei, and medullary raphe. In the pons and midbrain, retrogradely-labeled neurons of the same phenotypes were found in the Kölliker-Fuse and parabrachial nuclei, periaqueductal gray, pedunculopontine nucleus (PPT) and laterodorsal tegmentum (LDT). We also identified somatostatin-expressing neurons in the preBötC and PHOX2B immunopositive cells in the pFV , NTS, and part of the RF. Surprisingly, we found no catecholaminergic neurons in the NTS, A5 or Locus Coeruleus, no serotoninergic raphe neurons nor any cholinergic neurons in the PPT and LDT that projected to the pFL . Our results indicate that pFL neurons receive extensive excitatory and inhibitory inputs from several respiratory and nonrespiratory related brainstem regions that could contribute to the complex modulation of the conditional pFL oscillator for active expiration.

Brain-wide Mapping of Mono-synaptic Afferents to Different Cell Types in the Laterodorsal Tegmentum.

The laterodorsal tegmentum (LDT) is a brain structure involved in distinct behaviors including arousal, reward, and innate fear. How environmental stimuli and top-down control from high-order sensory and limbic cortical areas converge and coordinate in this region to modulate diverse behavioral outputs remains unclear. Using a modified rabies virus, we applied monosynaptic retrograde tracing to the whole brain to examine the LDT cell type specific upstream nuclei. The LDT received very strong midbrain and hindbrain afferents and moderate cortical and hypothalamic innervation but weak connections to the thalamus. The main projection neurons from cortical areas were restricted to the limbic lobe, including the ventral orbital cortex (VO), prelimbic, and cingulate cortices. Although different cell populations received qualitatively similar inputs, primarily via afferents from the periaqueductal gray area, superior colliculus, and the LDT itself, parvalbumin-positive (PV+) GABAergic cells received preferential projections from local LDT neurons. With regard to the different subtypes of GABAergic cells, a considerable number of nuclei, including those of the ventral tegmental area, central amygdaloid nucleus, and VO, made significantly greater inputs to somatostatin-positive cells than to PV+ cells. Diverse inputs to the LDT on a system-wide level were revealed.

Afferent connections of the dorsal, perigenual, and subgenual anterior cingulate cortices of the monkey: Amygdalar inputs and intrinsic connections.

The anterior cingulate cortex (ACC) is crucial for emotional processing, and its abnormal activities contributes to mood disorders. The ACC is divided into three subregions: the dorsal ACC (dACC), perigenual ACC (pgACC), and subgenual ACC (sgACC). Although these regions have been implicated in emotional processing, the dACC is more involved in cognitive functions, while the other two regions are important in the pathophysiology underlying mood disorders. Recent studies have suggested that the sgACC and pgACC exhibit opposite emotion-related activity patterns and that an interaction of the ACC with the amygdala is crucial for emotion-related ACC functions. Here, we injected neuronal tracers into the sgACC, pgACC, and dACC of macaques and quantitatively compared the distributions of the retrogradely labeled neurons in the amygdalar nuclei. For both the dACC and pgACC, about 90% of the labeled neurons were found in the basal nucleus, about 10% were in the accessory basal nucleus, and the lateral nucleus had almost no neuronal labeling. However, after sgACC injections, nearly half of the labeled neurons were found in the accessory basal nucleus, and a moderate number of labeled neurons were found in the lateral nucleus. These differences in amygdalar inputs might underlie the functional differences in the sgACC and pgACC. Moreover, after tracer injections in the sgACC, labeled neurons were observed in the pgACC and not the dACC, suggesting that the pgACC directly influences the activity of the sgACC.

A pathway from midcingulate cortex to posterior insula gates nociceptive hypersensitivity.

The identity of cortical circuits mediating nociception and pain is largely unclear. The cingulate cortex is consistently activated during pain, but the functional specificity of cingulate divisions, the roles at distinct temporal phases of central plasticity and the underlying circuitry are unknown. Here we show in mice that the midcingulate division of the cingulate cortex (MCC) does not mediate acute pain sensation and pain affect, but gates sensory hypersensitivity by acting in a wide cortical and subcortical network. Within this complex network, we identified an afferent MCC-posterior insula pathway that can induce and maintain nociceptive hypersensitivity in the absence of conditioned peripheral noxious drive. This facilitation of nociception is brought about by recruitment of descending serotonergic facilitatory projections to the spinal cord. These results have implications for our understanding of neuronal mechanisms facilitating the transition from acute to long-lasting pain.

Anterior and posterior parts of the rat ventral tegmental area and the rostromedial tegmental nucleus receive topographically distinct afferents from the lateral habenular complex.

That activation of the reward system involves increased activity of dopaminergic (DA) neurons in the ventral tegmental area (VTA) is widely accepted. In contrast, the lateral habenular complex (LHb), which is known as the center of the anti-reward system, directly and indirectly inhibits DA neurons in the VTA. The VTA, however, is not a homogenous entity. Instead, it displays major functional differences between its anterior (aVTA) and posterior (pVTA) regions. It is not precisely known, whether habenular input to the aVTA, pVTA, and the newly recognized rostromedial tegmental nucleus (RMTg) are similarly or differently organized. Consequently, the present investigation addressed the connections between LHb and aVTA, pVTA, and RMTg using retrograde and anterograde tracing techniques in the rat. Our experiments disclosed strictly reciprocal and conspicuously focal interconnections between LHbM (LHbMPc/LHbMC) and PN, as well as between RLi and LHbLO. In addition, we found that LHb inputs to the aVTA are dorsoventrally ordered. Dorsal parts of the aVTA receive afferents from LHbL and LHbM, whereas ventral parts of the aVTA are preferentially targeted by the LHbM. LHb afferents to the pVTA are distinct from those to the RMTg, given that the RMTg is primarily innervated from the LHbL, whereas pVTA receives afferents from LHbM and LHbL. These data indicate the existence of two separate pathways from the LHb to the VTA, a direct and an indirect one, which may subserve distinct biological functions.

Afferent and efferent connections of the interpeduncular nucleus with special reference to circuits involving the habenula and raphe nuclei.

The habenula is an epithalamic structure differentiated into two nuclear complexes, medial (MHb) and lateral habenula (LHb). Recently, MHb together with its primary target, the interpeduncular nucleus (IP), have been identified as major players in mediating the aversive effects of nicotine. However, structures downstream of the MHb-IP axis, including the median (MnR) and caudal dorsal raphe nucleus (DRC), may contribute to the behavioral effects of nicotine. The afferent and efferent connections of the IP have hitherto not been systematically investigated with sensitive tracers. Thus, we placed injections of retrograde or anterograde tracers into different IP subdivisions or the MnR and additionally examined the transmitter phenotype of major IP and MnR afferents by combining retrograde tract tracing with immunofluorescence and in situ hybridization techniques. Besides receiving inputs from MHb and also LHb, we found that IP is reciprocally interconnected mainly with midline structures, including the MnR/DRC, nucleus incertus, supramammillary nucleus, septum, and laterodorsal tegmental nucleus. The bidirectional connections between IP and MnR proved to be primarily GABAergic. Regarding a possible topography of IP outputs, all IP subnuclei gave rise to descending projections, whereas major ascending projections, including focal projections to ventral hippocampus, ventrolateral septum, and LHb originated from the dorsocaudal IP. Our findings indicate that IP is closely associated to a distributed network of midline structures that modulate hippocampal theta activity and forms a node linking MHb and LHb with this network, and the hippocampus. Moreover, they support a cardinal role of GABAergic IP/MnR interconnections in the behavioral response to nicotine.

Chemical anatomy of pallidal afferents in primates.

Neurons of the globus pallidus receive massive inputs from the striatum and the subthalamic nucleus, but their activity, as well as those of their striatal and subthalamic inputs, are modulated by brainstem afferents. These include serotonin (5-HT) projections from the dorsal raphe nucleus, cholinergic (ACh) inputs from the pedunculopontine tegmental nucleus, and dopamine (DA) afferents from the substantia nigra pars compacta. This review summarizes our recent findings on the distribution, quantitative and ultrastructural aspects of pallidal 5-HT, ACh and DA innervations. These results have led to the elaboration of a new model of the pallidal neuron based on a precise knowledge of the hierarchy and chemical features of the various synaptic inputs. The dense 5-HT, ACh and DA innervations disclosed in the associative and limbic pallidal territories suggest that these brainstem inputs contribute principally to the planification of motor behaviors and the regulation of attention and mood. Although 5-HT, ACh and DA inputs were found to modulate pallidal neurons and their afferents mainly through asynaptic (volume) transmission, genuine synaptic contacts occur between these chemospecific axon varicosities and pallidal dendrites, revealing that these brainstem projections have a direct access to pallidal neurons, in addition to their indirect input through the striatum and subthalamic nucleus. Altogether, these findings reveal that the brainstem 5-HT, ACh and DA pallidal afferents act in concert with the more robust GABAergic inhibitory striatopallidal and glutamatergic excitatory subthalamopallidal inputs. We hypothesize that a fragile equilibrium between forebrain and brainstem pallidal afferents plays a key role in the functional organization of the primate basal ganglia, in both health and disease.

Somatotopic map of the active electrosensory sense in the midbrain of the mormyrid Gnathonemus petersii.

In many vertebrates parallel processing in topographically ordered maps is essential for efficient sensory processing. In the active electrosensory pathway of mormyrids afferent input is processed in two parallel somatotopically ordered hindbrain maps of the electrosensory lateral line lobe (ELL), the dorsolateral zone (DLZ), and the medial zone (MZ). Here phase and amplitude modulations of the self-generated electric field were processed separately. Behavioral data indicates that this information must be merged for the sensory system to categorically distinguish capacitive and resistive properties of objects. While projections between both zones of the ELL have been found, the available physiological data suggests that this merging takes place in the midbrain torus semicircularis (TS). Previous anatomical data indicate that the detailed somatotopic representation present in the ELL is lost in the nucleus lateralis (NL) of the TS, while a rough rostrocaudal mapping is maintained. In our study we investigated the projections from the hindbrain to the midbrain in more detail, using tracer injections. Our data reveals that afferents from both maps of the ELL terminate in a detailed somatotopic manner within the midbrain NL. Furthermore, we provide data indicating that phase and amplitude information may indeed be processed jointly in the NL. J. Comp. Neurol. 524:2479-2491, 2016. © 2016 Wiley Periodicals, Inc.