Tracking of Fascicles of Cutaneous Nerves of Thigh: A Histological Study

Present study uncovers the secrets of internal morphology of femoral nerve branches namely, cutaneous trunk, subcutaneous trunks, saphenous, medial cutaneous and intermedius cutaneous nerves innervating the skin of anteromedial thigh at fascicular level. Therefore, the aim of the study is to track, correlate, interpret and identify the pathways of fascicles through histological slides. The femoral nerve and its branching points were calibrated in distances from inguinal ligament. These trunks and nerves of a cadaver were processed for histological slides staining with haematoxylin and eosin. The fascicles in the histological slides were identified, tracked, correlated and interpreted from cranial most slide to the last terminal slides of these nerves and trunks. The correlation of the pathways of fascicles revealed that these fascicles are continuous, consistent and traceable interrupted by split, fusion and multiplexing. Femoral nerve branches/fascicles/nerve fibres if damaged, impair the sensation of corresponding area of skin of anteromedial thigh creating helm of neurological complications. Hence the injured fascicles can be repaired with the help of identification and correlation of fascicular pathways carried out in this study with least invasion. The findings of present study will be of paramount importance for intraoperative stimulation to diagnose and identify the fascicle for microneurosurgical repair/graft/regenerate/neurotisation in the cutaneous branches of femoral nerve at fascicular level.


Introduction
There are many variations of femoral nerve and these have been classified by Singh et al. [1]. A femoral nerve cropped from a cadaver was type II of classification of Singh et al. [1]. This femoral nerve bifurcated into muscular and cutaneous trunks at one centimetre below the inguinal ligament. The cutaneous trunk further splits into sub-cutaneous trunk of thigh and the saphenous (S) nerve. The subcutaneous trunk, then, bifurcates into intermedius cutaneous (IC) and medial cutaneous (MC) nerves of the thigh. The group of afferent fascicles of S, IC and MC supply skin of anteromedial thigh.
Though the injuries to the IC and MC nerves have hardly been reported yet few cases of pain and paraesthesia over the anterior and medial aspects of thigh, as a result of engagement of IC and MC nerves of the thigh, are described. However, sensory loss on the medial side of the thigh, leg and foot up to the ball of the great toe because of engagement of the saphenous nerve through iatrogenic lapses or otherwise are well reported. The outcome of injuries may not be fatal or produce unbearably serious signs and symptoms so the patients may not be opting for costly neurosurgical diagnosis (MRI) for detection of location and degree of injury and procedures.
The neuro-therapy of neuropathological morbidity requires accurate diagnosis and treatment. There is very limited scope of investigating location and identification of injured fascicle or nerve fibres under current knowledge of internal morphology of nerve. Though Chhabra et al. claims that location and degree of injury can be identified through MR advanced neurography [2] yet it has its own limitations regarding resolution and image defects. Therefore, a micro-anatomic study has been planned to track fascicles in histological slides of subcutaneous trunk, S, IC and MC for improving identification of injured fascicle, its location and degree of injury for diagnosis and imagery interpretation together with non-invasive neurosurgical repair, grafting and regeneration of injured nerve fibres.

Tracking and correlation of fascicles
A24 slide was prepared from the femoral nerve just above the inguinal ligament. This femoral nerve just below the inguinal ligament bifurcated into muscular and cutaneous trunks. Cutaneous trunk then bifurcated into saphenous nerve and subcutaneous trunk which further divided into intermedius and medial cutaneous nerves. Saphenous, intermedius and medial cutaneous nerves innervate skin of anteromedial thigh. Histological slides of cutaneous, saphenous nerve, subcutaneous trunk, medial and intermediate cutaneous nerves were prepared and stained with haematoxylin and eosin. The fascicles in these nerves/trunks were identified, tracked, correlated and interpreted.

Naming scheme of fascicles
For deciphering the fascicles of individual nerves and to avoid confusing duplicate numbers, the name of the fascicles at the point of transformational processes was changed in sequential order with prefix from CF of composite fascicles in femoral nerve to CCF in cutaneous trunk and SCCFs in subcutaneous trunk extending it to SCF in S nerve, MCCFs and ICCFs in MC and IC respectively.

Tracking and correlation of fascicles in cutaneous trunk
The cutaneous trunk was cut into 6 pieces and six blocks C1, C2, C3, C4, C5 and C6 were prepared. From C1, 13 slides were processed. C1 13 was the cranial most slide of C1. Similarly variable number of slides were prepared from C2, C3, C4, C5 and C6.
In C24 (Figure 1). These fascicles continuously consistent up to slide C2 16, the cranial most slide of C2 block.
The fascicles in C2 16 were continuous, consistent and correctable up to slide C2 12. In slide C2 12, CF312 and CCF318 fused forming CCF320 and CF280 in slide C2 12 split into CCF321, 322 and 323 in C2 11 (Figure 2). C2 11 is traceable to C2 1 meaning that the fascicles which were present in slide C2 11 were continuing up to slide C2 1.

Tracking and correlation of fascicles in saphenous nerve
S nerve was cut into 6 pieces and six blocks S1, S2, S3, S4, S5 and S6 were prepared. Slides prepared from these six blocks were stained with haematoxylin and eosin and fascicles of S nerve were correlated starting from S1 block to S6 block.
The S nerve having CCFs 319, 320, 323, 326 and 327 in S1 1 emerged out after C6 1 (Figure 7). These CCFs were traceable from the slides of block S1 through S2 1 the caudal most slide of S2 block.
The fascicles are traceable between S6 11 and S6 7. Again reorganisation of fascicles is taking place from S6 7 to S6 6 (Figure 12). These fascicles in S6 6 were continuous, consistent and traceable up to slide S6 1. Infrapatellar branch emanated laterally from S nerve after S6 1.

Tracking and correlation of fascicles in subcutaneous trunk
Subcutaneous trunk was cut into 3 pieces and 3 blocks SCT1, SCT2 and SCT3 were prepared. Subcutaneous trunk bifurcated into MC and IC. One block of   MC,MC1 and two blocks of IC, IC1 and IC2 were prepared and slides from aforementioned blocks were stained with haematoxylin and eosin and fascicles were correlated as elaborated below: The CFs 303 and 304 constituted subcutaneous trunk which was separated laterally from cutaneous trunk after C6 1 (Figure 6). Then CF303 split into SCCF 305 and 306 in SCT1 1 slide (Figure 13). So the subcutaneous trunk now consists of fascicle CFs 304, SCCFs 305 and 306. After the slide SCT1 1, the CFs 304, SCCFs 305 and 306 were traceable from SCT1 1 up to SCT3 1.

Clinical significance of this study
No such study has been carried out involving sensory nerves of femoral nerve. If the fascicles of S, MC and IC are damaged, the communication of sensory information from the innervated area will be interrupted leading to aggravation of clinical problems. So it is not merely nerve where injury should be investigated rather injured fascicles should be targeted for diagnosis for complications which may occur anywhere in entire fascicular path from origin to point of innervation. The diagnosis of neural insults requires not only the location and degree of injury but also identification, isolation, orientation, directivity, and matching of shape and size of injured nerve CFs for planning surgical repair, grafting and regeneration [3].
The location and degree of injury is investigated by the high resolution MRI advanced neurography [2,4]. But this has its own limitations of recording and interpretation. This generates uncertainty in diagnosis and thereby in treatment. Thus the radiologists and neurosurgeons face the impediments of pinpointing the probable position of injury and identification of fascicles. Therefore, the imagery coupled with our internal morphological study together can refine the interpretation for identification of injured CF and location of injury. Methodically, it can be done by one to one correlation between images of transverse histological and high resolution MRI advanced neurographic sections at the same position from inguinal ligament. The distance of location of injured fascicle from inguinal ligament may be computed in MRI neurography and then the calibrated histological sections of cutaneous, subcutaneous trunks, S, MC and IC nerves at the same level may be compared and examined for confirmation of identified injured fascicle. After identification of injured fascicle, the idea of shape, size, location and orientation can also be derived from histological slides for matching, alignment and directivity of nerve fibres for repair and grafting.

Personal communication
The neurosurgery at fascicular level is currently uncommon however, with upcoming science and technology in future, present study will be highly useful for neurosurgeons. The study will help in carrying out less invasive surgery as stimulation of identified injured fascicles will not involve other fascicles which in case of nerve stimulation may be stimulated causing discomfort to the patient.