Animal Molecular Breeding 2016, Vol.6, No.4, 1-10
2
Work done on the developmental aspects of avian brain include those by Davey and Tickle (2007) on the brain
development of chicken, Salami (2009) skeletal development of guinea fowl, Gossomji (2014) on the
development of GIT in guinea fowl and Serdar and Emrah (2010) cortical development of the chicken cerebellum.
However, basic information on the structural development of the cerebellum of the Grey breasted helmeted guinea
fowl is not currently available, hence the need for this investigation.
2 Materials and Methods
2.1 Experimental eggs
Seventy (70) fertilized guinea fowl eggs were purchased from National Veterinary Research Institution (NVRI)
Vom, Jos, Plateau State, Nigeria. The eggs were transported to a hatchery in Jos and incubated using their standard
incubation guide. During incubation, the eggs were turned regularly (minimum of three times) each day for the
first 24 days.
Fifty four (54) eggs (two eggs per day) for pre-hatch study were collected daily from day one of incubation up to
day twenty eight which is the final day of incubation. An opening was made on the large air space area and the
entire egg dropped into a labeled container of 10% buffered formalin for proper fixing. For post-hatch study, brain
samples from hatched keets were collected each week for eight weeks and also fixed in 10% buffered formalin.
2.2 Extraction of embryo from egg shell
This was done at pre-hatch using a scalpel blade and clean transparent dish. The blunt side of the scalpel blade
was used with the egg held on the palm, and a gentle tap was made on the egg until a crack was formed. Then, the
crack was gently widened manually and the embryo collected in a transparent dish.
2.3 Extraction of brain
At pre-hatch, because the entire skull is soft and pliable, scapel blade and rat tooth forceps were used for
extraction of the brain. At post-hatch, the keets were euthanized using Nembutal at 40 mg per body weight.
Immediately, decapitation was made and the heads fixed in 10% buffered formalin for 3–5 days. After ensuring
proper fixation, a dissection was made at the angle of the beak up to the level of the occipital bone. The upper
portion of the dissected area is pulled off gradually using the rat tooth forceps until the entire brain is exposed.
The cranial nerves were severed to ease the lifting of the brain from the cranium
.
The extracted brains were fixed
in Bouin’s solution comprising 75 ml picric acid, 25 ml formaldehyde and 5 ml glacial acetic acid.
2.4 Separation of the cerebellum
The cerebellum is located on the dorsal portion of the brainstem, with its peduncles: the restiform body connects
to the medulla, the brachium pontis connects the cerebellum to the Pons and the brachium conjunctivum connects
the cerebellum to the midbrain. The peduncles were severed using a scalpel blade to expose the entire brainstem.
2.5 Gross
Thirty five (27 for pre-hatch and 8 for post-hatch) brain samples were used. Photographs of the dorsal and ventral
aspects of the brain were taken using digital handheld Microscope (Magnification 1000x, 5x Zoom, 3D stand high
speed DSP).
2.6 Histological techniques
Thirty five (27 for pre-hatch and 8 for post-hatch) brain samples were used. Fixed brain samples were washed
using tap water and dehydrated through ascending grades of alcohol (70%, 80%, 90%, 95%, 100%), within the
interval of three (3) hours each, cleared in xylene for two hours and embedded in liquid paraffin at 50
o
C as
described by Kiernan (1990).
Serial transverse sections of 5 µ were made using Jung rotary microtome (Model 42339, Berlin, Germany).
Sections were mounted on glass slides and allowed to dry; were deparaffinized, stained, hydrated and cover
