Tahsinul Haque

Corticofugal projections to trigeminal motoneurons innervating antagonistic jaw muscles in rats as demonstrated by anterograde and retrograde tract tracing

Little is known about the organization of corticofugal projections controlling antagonistic jaw muscles. To address this issue, we employed retrograde (Fluorogold; FG) and anterograde (biotinylated dextran amine; BDA) tracing techniques in rats. Three groups of premotoneurons were identified by injecting FG into the jaw‐closing (JC) and ‐opening (JO) subdivisions of the trigeminal motor nucleus (Vmo). These were 1) the intertrigeminal region (Vint) and principal trigeminal sensory nucleus for JC nucleus; 2) the reticular region medial to JO nucleus (RmJO) for JO nucleus; and 3) the parabrachial (Pb) and supratrigeminal (Vsup) nuclei, reticular regions medial and ventral to JC nucleus, rostrodorsomedial oralis (Vor), and juxtatrigeminal region (Vjuxt) containing a mixture of premotoneurons to both the nuclei. Subsequently, FG was injected into the representative premotoneuron structures. The JC and JO premotoneurons received main afferents from the lateral and medial agranular fields of motor cortex (Agl and Agm), respectively, whereas afferents to the nuclei with both JC and JO premotoneurons arose from Agl also and from primary somatosensory cortex (S1). Finally, BDA was injected into each of the three cortical areas representing the premotoneuron structures to complement the FG data. The Agl and Agm projected to reticular regions around the Vmo, whereas the Pb, Vsup, Vor, and Vjuxt received input from Agl. The S1 projected to the trigeminal sensory nuclei as well as to the Pb, Vsup, and Vjuxt. These results suggest that corticofugal projections to Vmo via premotoneuron structures consist of multiple pathways, which influence distinct patterns of jaw movements. J. Comp. Neurol. 514:368–386, 2009.

Projections from the insular cortex to pain-receptive trigeminal caudal subnucleus (medullary dorsal horn) and other lower brainstem areas in rats.

This study examined the projections from the rat insular cortex (Ins) to lower brainstem areas which are possibly involved in orofacial pain processing. We first examined distributions of Ins neurons projecting directly to the trigeminal caudal subnucleus (Vc, medullary dorsal horn) and oral subnucleus (Vo) which are known to receive orofacial nociceptive inputs. After injections of a retrograde tracer, Fluorogold (FG), into the medial part and lateral part of laminae I/II of Vc, many neurons were labeled bilaterally with a contralateral predominance in the rostral level of granular Ins (GI) and dysgranular Ins (DI) and the caudal level of GI/DI, respectively, but none in the agranular Ins (AI). After FG injections into laminae III-V of Vc, no Ins neurons were labeled. After FG injections into the Vo, many neurons were labeled bilaterally with a contralateral predominance in the rostral and caudal GI/DI, but none in the AI. We then examined descending projections from the GI/DI to the lower brainstem. After injections of an anterograde tracer, biotinylated dextranamine (BDA), into the rostral GI/DI, many BDA-labeled axons and terminals were seen bilaterally with a contralateral predominance in the medial part of laminae I/II of Vc, dorsomedial Vo, juxtatrigeminal region, rostral ventromedial medulla (RVM), and nucleus of the solitary tract, and with an ipsilateral predominance in the parabrachial nucleus (Pb), Kölliker-Fuse nucleus (KF) and trigeminal mesencephalic nucleus. After BDA injections into the caudal GI/DI, they were seen bilaterally with a contralateral predominance in the lateral part of laminae I/II of Vc, ventrolateral Vo, juxtatrigeminal region and RVM, and with an ipsilateral dominance in the lateral zone (PAGl) of periaqueductal gray, Pb and KF. These results suggest that orofacial nociceptive processing of Vc and Vo neurons may be regulated by GI/DI directly or indirectly through brainstem nuclei such as PAGl, Pb, KF and RVM.

Somatotopic direct projections from orofacial areas of primary somatosensory cortex to pons and medulla, especially to trigeminal sensory nuclear complex, in rats

The primary somatosensory cortex (S1) projects to the thalamus and brainstem somatosensory nuclei and modulates somatosensory information ascending to the S1 itself. However, the projections from the S1 to the brainstem second-order somatosensory neuron pools have not been fully studied. To address this in rats, we first revealed the somatotopic representation of orofacial areas in the S1 by recording cortical surface potentials evoked by stimulation of the lingual, mental, infraorbital, and frontal nerves. We then examined the morphology of descending projections from the electrophysiologically defined orofacial S1 areas to the pons and medulla after injections of an anterograde tracer, biotinylated dextranamine (BDA), into the orofacial S1 areas. BDA-labeled axon terminals were seen mostly in the trigeminal sensory nuclear complex (TSNC) and had a strong contralateral predominance. They also showed a somatotopic arrangement in dorsoventral and superficial-deep directions within almost all rostrocaudal TSNC levels, and in a rostrocaudal direction within the trigeminal caudal subnucleus. In the principal nucleus (Vp) or oral subnucleus (Vo) of TSNC, the BDA-labeled axon terminals showed a somatotopic arrangement closely matched to that of the electrophysiologically defined projection sites of orofacial primary afferents; these projection sites were marked by injections of a retrograde tracer, Fluorogold (FG), into the Vp or Vo. The FG injections labeled a large number of S1 neurons, with a strong contralateral predominance, in a somatotopic manner, which corresponded to that presented in the electrophysiologically defined orofacial S1 areas. The present results suggest that the orofacial S1 projections to somatotopically matched regions of trigeminal second-order somatosensory neuron pools may allow the orofacial S1 to accurately modulate orofacial somatosensory transmission to higher brain centers including the orofacial S1 itself.

Thalamic afferent and efferent connectivity to cerebral cortical areas with direct projections to identified subgroups of trigeminal premotoneurons in the rat.

The roles of supramedullary brain mechanisms involved in the control of jaw movements are not fully understood. To address this issue, a series of retrograde (Fluorogold, FG) and anterograde (biotinylated dextran amine, BDA) tract-tracing studies were done in rats. At first, we identified projection patterns from defined sensorimotor cortical areas to subgroups of trigeminal premotoneurons that are located in defined brainstem areas. Focal injections of FG into these brainstem areas revealed that the rostralmost part of lateral agranular cortex (rmost-Agl), the rostralmost part of medial agranular cortex (rmost-Agm), and the rostralmost part of primary somatosensory cortex (rmost-S1) preferentially project to brainstem areas containing jaw-closing premotoneurons, jaw-opening premotoneurons and a mixture of both types of premotoneurons, respectively. The thalamic reciprocal connectivities to rmost-Agl, rmost-Agm, and rmost-S1 were then investigated following cortical injections of FG or BDA. We found many retrogradely FG-labeled neurons and large numbers of axons and terminals labeled anterogradely with BDA in the dorsal thalamus mainly on the side ipsilateral to the injection sites. The rmost-Agl had strong connections with the ventral lateral nucleus (VL), ventromedial nucleus (VM), parafascicular nucleus, and posterior nucleus (Po); the rmost-Agm with the ventral anterior nucleus, VL, VM, central lateral nucleus, paracentral nucleus, central medial nucleus, mediodorsal nucleus and Po; and the rmost-S1 with the ventral posteromedial nucleus and Po. The present results suggest that the descending multiple pathways from the cerebral cortex to jaw-closing and jaw-opening premotoneurons have unique functional roles in jaw movement motor control.

Corticofugal direct projections to primary afferent neurons in the trigeminal mesencephalic nucleus of rats.

Little is known about projections from the cerebral cortex to the trigeminal mesencephalic nucleus (Vmes) which contains the cell bodies of primary sensory afferents innervating masticatory muscle spindles and periodontal ligaments of the teeth. To address this issue, we employed retrograde (Fluorogold, FG) and anterograde (biotinylated dextranamine, BDA) tracing techniques in the rat. After injections of FG into the Vmes, a large number of neurons were retrogradely labeled in the prefrontal cortex including the medial agranular cortex, anterior cingulate cortex, prelimbic cortex, infralimbic cortex, deep peduncular cortex and insular cortex; the labeling was bilateral, but with an ipsilateral predominance to the injection site. Almost no FG-labeled neurons were found in the somatic sensorimotor cortex. After BDA injections into the prefrontal cortex, anterogradely labeled axon fibers and boutons were distributed bilaterally in a topographic pattern within the Vmes, but with an ipsilateral predominance to the injection site. The rostral Vmes received more preferential projections from the medial agranular cortex, while the deep peduncular cortex and insular cortex projected more preferentially to the caudal Vmes. Several BDA-labeled axonal boutons made close associations (possible synaptic contacts) with the cell bodies of Vmes neurons. The present results have revealed the direct projections from the prefrontal cortex to the primary sensory neurons in the Vmes and their unique features, suggesting that deep sensory inputs conveyed by the Vmes neurons from masticatory muscle spindles and periodontal ligaments are regulated with specific biological significance in terms of the descending control by the cerebral cortex.

SOMATOTOPIC DIRECT PROJECTIONS FROM OROFACIAL AREAS OF SECONDARY SOMATOSENSORY CORTEX TO TRIGEMINAL SENSORY NUCLEAR COMPLEX IN RATS

Little is known about the projections from the orofacial areas of the secondary somatosensory cortex (S2) to the pons and medulla including the second-order somatosensory neuron pools. To address this in rats, we first examined the distribution of S2 neurons projecting to the trigeminal principal nucleus (Vp) or oral subnucleus (Vo) of the trigeminal sensory nuclear complex (TSNC) after injections of a retrograde tracer, Fluorogold (FG), into five regions in the Vp/Vo which were responsive to stimulation of trigeminal nerves innervating the orofacial tissues. A large number of FG-labeled neurons were found with a somatotopic arrangement in the dorsal areas of S2 (orofacial S2 area). This somatotopic arrangement in the orofacial S2 area was shown to closely match that of the orofacial afferent inputs by recording cortical surface potentials evoked by stimulation of the trigeminal nerves. We then examined the morphology of descending projections from these electrophysiologically defined areas of the orofacial S2 to the pons and medulla after injections of an anterograde tracer, biotinylated dextranamine (BDA), into the areas. A large number of BDA-labeled axon fibers and terminals were seen only in some of the second-order somatosensory neuron pools, most notably in the contralateral TSNC, although the labeled terminals were not seen in certain rostrocaudal levels of the contralateral TSNC including the rostrocaudal middle level of the trigeminal interpolar subnucleus. The projections to the TSNC showed somatotopic arrangements in dorsoventral, superficial-deep and rostrocaudal directions. The somatotopic arrangements in the Vp/Vo closely matched those of the electrophysiologically defined central projection sites of the orofacial trigeminal afferents in the TSNC. The present results suggest that the orofacial S2 projects selectively to certain rostrocaudal levels of the contralateral TSNC, and the projections may allow the orofacial S2 to accurately modulate orofacial somatosensory transmission to higher brain centers including the orofacial S2 itself.

Distribution of premotoneurons for jaw-closing and jaw-opening motor nucleus receiving contacts from axon terminals of primary somatosensory cortical neurons in rats

To clarify features of direct projections from the primary somatosensory cortex (S1) to premotoneurons for the jaw-closing (JC) and jaw-opening (JO) components of the trigeminal motor nucleus, biotinylated dextranamine (BDA) and Fluorogold (FG) were used as the anterograde and retrograde tracers. The BDA and FG injections were made in the S1 and the JC or JO component, respectively, in rats. The distribution of FG-labeled JC and JO premotoneurons receiving contact(s) from BDA-labeled axon terminals of S1 neurons was quantitatively examined; the contacts were identified microscopically by using a X100 oil immersion objective. The largest and second largest numbers of JC and JO premotoneurons with contact(s) were found in the lateral reticular formation at the levels of the caudal pons and the medulla oblongata (cpmLRt) and trigeminal oral nucleus (Vo) bilaterally, and they comprised about 80% of the total premotoneurons with contact(s). The percentage of premotoneurons with contact(s) was higher in the Vo than in the cpmLRt for both JC and JO premotoneurons. Most of the JC or JO premotoneurons found in the nucleus of the solitary tract, inter- and supratrigeminal regions, mesencephalic trigeminal nucleus, parabrachial nucleus and reticular formation medial to the JO component of the trigeminal motor nucleus hardly received contact(s) from S1 neurons. This suggests that the contribution of S1 to the control of jaw movements is mediated via JC and JO premotoneurons located primarily in the cpmLRt and Vo areas of the brainstem.

Projections from the dorsal peduncular cortex to the trigeminal subnucleus caudalis (medullary dorsal horn) and other lower brainstem areas in rats.

This study has revealed direct projections from the dorsal peduncular cortex (DP) in the medial prefrontal cortex (mPfC) to the trigeminal brainstem sensory nuclear complex and other lower brainstem areas in rats. We first examined the distribution of mPfC neurons projecting directly to the medullary dorsal horn (trigeminal subnucleus caudalis [Vc]) and trigeminal subnucleus oralis (Vo) which are known to receive direct projections from the lateral prefrontal cortex (insular cortex). After injections of the retrograde tracer Fluorogold (FG) into the rostro-dorsomedial part of laminae I/II of Vc (rdm-I/II-Vc), many neurons were labeled bilaterally (with an ipsilateral predominance) in the rostrocaudal middle level of DP (mid-DP) and not in other mPfC areas. After FG injections into the lateral and caudal parts of laminae I/II of Vc, or the Vo, no neurons were labeled in the mPfC. We then examined projections from the mid-DP by using the anterograde tracer biotinylated dextranamine (BDA). After BDA injections into the mid-DP, many axons and terminals were labeled bilaterally (with an ipsilateral predominance) in the rdm-I/II-Vc, periaqueductal gray and solitary tract nucleus, and ipsilaterally in the parabrachial nucleus and trigeminal mesencephalic nucleus. In addition, the connections of the mid-DP with the insular cortex were examined. Many BDA-labeled axons and terminals from the mid-DP were also found ipsilaterally in the caudalmost level of the granular and dysgranular insular cortex (GI/DI). After BDA injections into the caudalmost GI/DI, many axons and terminals were labeled ipsilaterally in the mid-DP. The projections from the mid-DP to the rdm-I/II-Vc and other brainstem nuclei suggest that mid-DP neurons may regulate intraoral and perioral sensory processing (including nociceptive processing) of rdm-I/II-Vc neurons directly or indirectly through the brainstem nuclei. The reciprocal connections between the mid-DP and caudalmost GI/DI suggest that this regulation may involve mid-DP interactions with the caudalmost GI/DI neurons.

Jaw-opening and -closing premotoneurons in the nucleus of the solitary tract making contacts with laryngeal and pharyngeal afferent terminals in rats.

This study clarified the neural mechanisms underlying jaw movements in pharyngolaryngeal reflexes such as swallowing in rats. After retrograde tracer injections into the ventromedial division (Vmovm) of the trigeminal motor nucleus (Vmo) containing jaw-opening (JO) motoneurons or into the dorsolateral division (Vmodl) of Vmo containing jaw-closing (JC) motoneurons, JO and JC premotoneurons were labeled with an ipsilateral predominance in the medial and intermediate subnuclei of the rostrocaudal middle two-thirds of the nucleus of the solitary tract (Sol); JC premotoneurons were also in the lateral subnucleus of Sol. After anterograde tracer injections into the Sol, axons were labeled with an ipsilateral predominance in the Vmovm and Vmodl, prominently in the ipsilateral Vmovm. After transganglionic tracer applications to the superior laryngeal nerve (SLN) or the cervical trunk of the glossopharyngeal nerve (GpN-ct), labeled afferents were seen in the medial, intermediate, lateral and interstitial subnuclei of Sol at the rostral three-fourths of Sol, indicating considerable overlap with the JO and JC premotoneurons in the Sol. Double labeling experiments demonstrated contacts between the afferent terminals and the JO and JC premotoneurons. The present study has for the first time revealed the differential distribution of JO and JC premotoneurons in the Sol and features of their projections from the Sol, as well as their connections with SLN and GpN-ct afferent inputs. The JO and JC premotoneurons in the Sol may play an important role in generation and organization of jaw movements in pharyngolaryngeal reflexes evoked by SLN and GpN-ct inputs, such as swallowing.

Thalamo-insular pathway conveying orofacial muscle proprioception in the rat

Little is known about how proprioceptive signals arising from muscles reach to higher brain regions such as the cerebral cortex. We have recently shown that a particular thalamic region, the caudo-ventromedial edge (VPMcvm) of ventral posteromedial thalamic nucleus (VPM), receives the proprioceptive signals from jaw-closing muscle spindles (JCMSs) in rats. In this study, we further addressed how the orofacial thalamic inputs from the JCMSs were transmitted from the thalamus (VPMcvm) to the cerebral cortex in rats. Injections of a retrograde and anterograde neuronal tracer, wheat-germ agglutinin-conjugated horseradish peroxidase (WGA-HRP), into the VPMcvm demonstrated that the thalamic pathway terminated mainly in a rostrocaudally narrow area in the dorsal part of granular insular cortex rostroventrally adjacent to the rostralmost part of the secondary somatosensory cortex (dGIrvs2). We also electrophysiologically confirmed that the dGIrvs2 received the proprioceptive inputs from JCMSs. To support the anatomical evidence of the VPMcvm-dGIrvs2 pathway, injections of a retrograde neuronal tracer Fluorogold into the dGIrvs2 demonstrated that the thalamic neurons projecting to the dGIrvs2 were confined in the VPMcvm and the parvicellular part of ventral posterior nucleus. In contrast, WGA-HRP injections into the lingual nerve area of core VPM demonstrated that axon terminals were mainly labeled in the core regions of the primary and secondary somatosensory cortices, which were far from the dGIrvs2. These results suggest that the dGIrvs2 is a specialized cortical region receiving the orofacial proprioceptive inputs. Functional contribution of the revealed JCMSs-VPMcvm-dGIrvs2 pathway to Tourette syndrome is also discussed.